Quantitative proteomic analysis of distinctmammalian Mediator complexes usingnormalized spectral abundance factorsAndrew C. Paoletti*, Tari J. Parmely*, Chieri Tomomori-Sato*, Shigeo Sato*, Dongxiao Zhu*, Ronald C. Conaway*,Joan Weliky Conaway*, Laurence Florens*, and Michael P. Washburn*

*Stowers Institute for Medical Research, Kansas City, MO 64110; Department of Biochemistry and Molecular Biology, Kansas University Medical Center,Kansas City, KS 66160; and Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73190

Edited by Roger D. Kornberg, Stanford University School of Medicine, Stanford, CA, and approved October 20, 2006 (received for review July 26, 2006)

Components of multiprotein complexes are routinely determinedby using proteomic approaches. However, this information lacksfunctional content except when new complex members are iden-tified. To analyze quantitatively the abundance of proteins inhuman Mediator we used normalized spectral abundance factorsgenerated from shotgun proteomics data sets. With this approachwe define a common core of mammalian Mediator subunits sharedby alternative forms that variably associate with the kinase moduleand RNA polymerase (pol) II. Although each version of affinity-purified Mediator contained some kinase module and RNA pol II,Mediator purified through F-Med26 contained the most RNA pol IIand the least kinase module as demonstrated by the normalizedspectral abundance factor approach. The distinct forms of Mediatorwere functionally characterized by using a transcriptional activityassay, where F-Med26 Mediator/RNA pol II was the most active.This method of protein complex visualization has important im-plications for the analysis of multiprotein complexes and assemblyof protein interaction networks.

multidimensional protein identification technology proteomics spectrum counting mass spectrometry

S ince its discovery in yeast (1, 2) and subsequent isolation andinitial characterizations in human cells (3), the transcrip-tional coactivator complex Mediator has been the subject ofnumerous studies to characterize both its composition andfunction. Researchers have used a combination of immunopre-cipitation (314), ion exchange/size-exclusion chromatography(8, 10, 1519), glycerol gradient (6, 7, 12, 13, 19, 20), SDS/PAGE/silver stain/Western blot analysis (6, 7, 9, 10, 15, 18, 21), MS (9,16, 19, 21, 22), and ChIP (21, 23) to determine the compositionofMediator complexes. Varying in size and subunit composition,the larger complexes (thyroid hormone receptor-associated pro-tein complex, SRB/Med-containing cofactor complex, negativeregulator of activated transcription, vitamin D receptor inter-acting protein complex, and activator recruited cofactor) rangedfrom 1 to 2 MDa and were composed of 30 subunits (3, 916,21, 22, 24), whereas the smaller complexes (cofactor required forSp1, positive cofactor 2, and positive cofactor 4) ranged from500 to 700 kDa and were composed of 917 subunits (57,1720, 25, 26). The main shared difference between large andsmall complexes appears to be the presence or absence of thekinase module. Furthermore, different complexes possesseddifferent levels of basal transcription, which could be accountedfor by several possibilities, including purification method (highionic strength may have washed away necessary proteins), theadding back of RNA polymerase (pol) II and basal transcriptionfactors, and testing only for activated transcription.Although these studies form the foundation for our present

understanding of Mediator, the relationship between differentforms ofMediator was unclear and the composition of each formwas unclear. The first attempt at standardizing Mediator com-plex purifications was carried out by using FLAG-tagged sub-

units of Mediator analyzed by multidimensional protein identi-fication technology (MudPIT) (27). This study definitivelyestablished a set of consensus Mediator subunits, determinedthat each of these subunits was present in almost every prepa-ration, identified new subunits (Cdk8L and Med13L), anddemonstrated that RNA pol II subunits were present in allpreparations (27). Nevertheless, our previous study provided noquantitative information, and the relative subunit stoichiome-tries and abundances were not determined.We therefore established a strategy to use MS as a means of

quantifying the relative abundance of all proteins present inMediator complexes. Although this study applies the sameanalytical means used to establish Mediator composition (27,28), it implements methods of data analysis that account forprotein size and variability between runs to normalize relativeprotein abundance between samples (termed NSAF values fornormalized spectral abundance factor) (29). NSAF is based onspectral counting, which has been demonstrated to be an effec-tive quantitative proteomics approach (3035). In spectralcounting, larger proteins would be expected to generate morepeptides and therefore more spectral counts than smaller pro-teins. Consequently, the number of spectral counts for eachprotein must be divided by the mass (33) or protein length, whichdefines the spectral abundance factor (SAF) (31). However, toaccurately account for run to run variation, individual SAFvalues must be normalized to one by dividing by the sum of allSAFs for proteins in the complex, resulting in the NSAF value(29). In a manner similar to microarray data analysis, NSAFsvalues are hence standardized across distinct preparations ofMediator to allow direct comparison between individual runs.The current quantitative analysis points out significant and

functionally relevant differences betweenMediator preparationspurified through different FLAG-tagged subunits: a commoncore of mammalian Mediator subunits is shared by alternativeforms that variably associate with the kinase module and RNApol II. Importantly, to substantiate this methodology as anaccurate and reliable method, not only to determine relativeprotein abundance, but also to predict complex activity, the MS

Author contributions: R.C.C., J.W.C., L.F., and M.P.W. designed research; A.C.P., T.J.P.,C.T.-S., and S.S. performed research; C.T.-S. and S.S. contributed new reagents/analytictools; A.C.P., T.J.P., D.Z., R.C.C., J.W.C., L.F., and M.P.W. analyzed data; and A.C.P., D.Z.,R.C.C., J.W.C., L.F., and M.P.W. wrote the paper.

The authors declare no conflict of interest.

This article is a PNAS direct submission.

Freely available online through the PNAS open access option.

Abbreviations: SAF, spectral abundance factor; NSAF, normalized SAF; MudPIT, multidi-mensional protein identification technology; pol, polymerase.

To whom correspondence should be addressed at: Stowers Institute for Medical Research,1000 East 50th Street, Kansas City, MO 64110. E-mail: mpw@stowers-institute.org.

This article contains supporting information online at www.pnas.org/cgi/content/full/0606379103/DC1.

2006 by The National Academy of Sciences of the USA

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data were verified by several orthogonal methods. We carriedout an AQUA experiment (36) using 15N Med9, and relativeprotein abundance estimated by NSAF was corroborated byWestern blot analysis to multiple proteins in different Mediatormodules andRNApol II, as well as by in vitro transcription assaysusing a purified basal transcription system (37). As opposed toprevious studies using spectral counting methods, in the currentarticle we demonstrate and verify the ability of the NSAFapproach to determine relative protein abundance in statisticallysupported structure/function analysis of multiprotein complexes.

ResultsMediator Core Subunits. Based on a combination of biochemical,genetic, and electron microscopy data, largely performed withthe Mediator complex from Saccharomyces cerevisiae, Mediatorsubunits have been assigned to several subassemblies, called thehead, middle, tail, and kinase modules (38). Mediator interactswith RNA pol II to form a larger complex that has been referredto as the RNA pol II holoenzyme (1, 4, 39). Because Mediatorproteins have been assigned to particular modules based largelyon studies of the S. cerevisiae complex (38), and because meta-zoan Mediator includes additional subunits not found in yeast(27), there are six Mediator subunits that have not yet beenlocalized to the head, middle, or tail regions.NSAF values measured for head, middle, tail, and unassigned

subunits in each of the four Mediator preparations are plottedin Fig. 1 AC. Regardless of the bait used to purify the

complexes, each core mediator protein appears to be present atlargely the same levels in each preparation. Statistically signif-icant exceptions included the baits Med26 and Med29, whichwere recovered at much higher levels in their own pull-downs,and Med30 in the F-Med28 pull-downs and Med31 in theF-Med10 preparations [Fig. 1 and supporting information (SI)].To validate the NSAF approach we carried out an AQUAexperiment (36) by adding various absolute amounts of recom-binant 15N Med9 purified from Escherichia coli into each of thefour Mediator preparations analyzed (Fig. 1B Inset). NSAFvalues for 15N Med9 showed a linear response over an 6-foldrange of abundance, whereas NSAF values for endogenousMed9 in each of the four Mediator preparations were notsignificantly different. To validate further the NSAF measure-ments we performed quantitative Western blotting using anti-bodies directed against Med17, Med8, and Med18. Both NSAFvalues (Fig. 1 AC) and Western blotting (Fig. 1D) demonstratethat the abundance of each of these components does not varyeven when different FLAG-tagged baits are used to purifyMediator. However, various levels are observed for differentproteins. For example, Med21 is overall the most abundantprotein component in the middle module, which suggests that itis present at more than one copy per Mediator complex or existsoutside of Mediator, either on its own or as part of anotherprotein complex present in the preparation. On the other end ofthe spectrum, Med25 is the least abundant of all core Mediatorcomponents regardless of the bait used to purify Mediator (Fig.

Fig. 1. NSAF analysis of human Mediator core components. NSAF values for Mediator components are plotted for each of the preparations purified throughF-Med26, which is bait 1 (B1), F-Med29 (B2), F-Med28 (B3), and F-Med10 (B4). (A) Head components of Mediator. (B) Middle and tail components. (C) Unlocalizedcomponents. Each column is the average of four independent analyses, and error bars represent one SD of the data. NSAFs that were statistically different (P0.05) from NSAF values measured in the other three preparations (SI) are designated with a vertical **. (B Inset) The endogenous Med9 NSAF data (solid bars)compared with a 15N Med9 standard curve (striped bars) where the absolute amount of purified 15N Med9 added to F-Med29, F-Med26, F-Med10, and F-Med28preparations was 0.128g, 0.113g, 0.042g, and 0.018g, respectively. (D) Quantitative Western blotting of the human Mediator components Med17, Med8,and Med18 from each of the affinity-purified Mediator preparations.

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1B). This NSAF result is in excellent agreement with previousstudies that demonstrated that Med25, which interacts withVP16, is easily dissociated from Mediator and is present atsubstoichiometric levels (11, 40).

Mediator Association with Kinase Module and RNA Pol II. The moststriking differences between Mediator complexes purifiedthrough different baits were in the relative amounts of kinasemodule (Fig. 2) and RNA pol II (Fig. 3) present in eachpreparation. The kinase module components Cdk8L, Med12,Med13, andMed13L were all significantly decreased inMediatorpurified through F-Med26 compared with Mediator purifiedthrough F-Med10, F-Med28, or F-Med29 (Fig. 2A and SI).Although NSAF values measured for the other two componentsof the kinase module, Cdk8 and Cyclin C, did not achieverigorous statistical significance, quantitative Western blottingrevealed that indeed Cdk8 was not detected and Cyclin C wasonly weakly detected in F-Med26 Mediator (Fig. 2B). On theother hand, the Mediator preparations purified through F-Med10, F-Med28, or F-Med29 all contained similar, and sub-stantially higher, levels of kinase module components as deter-mined both by the NSAF value (Fig. 2A), quantitative Westernblotting (Fig. 2B), and statistical analysis (see SI). Next, Medi-ator purified through different FLAG-tagged subunits alsocontained various amounts of RNA pol II (Fig. 3). As shown in

Fig. 3A, comparison of NSAF values for RNA pol II subunitsindicates that similar amounts of RNA pol II were recovered inF-Med10 and F-Med26 Mediator, whereas Mediator purifiedthrough F-Med28 and F-Med29 contained statistically signifi-cantly less RNA pol II. Again, this was validated with quanti-tative Western blotting against the RNA pol II subunits Rpb1and Rpb9, where F-Med26 Mediator had the most Rpb1 andRpb9 followed by F-Med10, with F-Med29 and F-Med28 prep-arations having the least RNA pol II subunits (Fig. 3B).

Fig. 2. NSAF analysis of the kinase module of human Mediator. (A) NSAFvalues for kinase module components are plotted for each of the baits used toaffinity-purify Mediator. Listed on the x axis is the bait protein. Above eachbait protein from left to right are Cdk8, Cdk8L, cyclin C, Med12, Med13, andMed13L. Each column is the average of four independent analyses, and errorbars represent one SD of the data. Because of sequence homology NSAF valuesfor Cdk8/Cdk8L are likely to be uniformly inflated because shared peptideswere used to calculate NSAFs. Statistically significant (P 0.05) average SDNSAF values for kinase module components in F-Med26 Mediator that werelower than NSAF values measured from the other three preparations areshown with vertical ** (see SI). (B) Quantitative Western blotting of the kinasemodule components Cdk8 and Cyclin C from each of the affinity-purifiedMediator preparations. The Med17 Western blot analysis is the same as shownin Fig. 1 and is provided as a frame of reference.

Fig. 3. NSAF analysis of RNA pol II components copurifying with humanMediator. (A) NSAF values for RNA pol II components are plotted for each ofthe baits used to purify Mediator. From left to right above each bait is theaverage NSAF value (with error bars) of Rpb1 to Rpb12 in numerical order.RNA pol II components in F-Med26 and F-Med10 purifications that werestatistically different (P 0.05) from all other preparations are represented bya vertical **, and RNA pol II components in F-Med26 and F-Med10 prepara-tions that were only significantly different from F-Med28 and F-Med29 prep-arations are represented by *. Rpb7 in F-Med10 was significantly differentfrom F-Med26 and F-Med29 but not F-Med28, and this is represented by (seeSI). (B) Quantitative Western blotting of the RNA pol II components Rpb1 andRpb9 from each of the affinity-purified Mediator preparations. The Med17Western blot analysis is the same as shown in Fig. 1 and is provided as a frameof reference. (C) Basal transcription performed on a naked DNA template withand without added RNA pol II. The amount of added eluate was normalizedto Med17 subunit as determined from Western blot analysis. Transcriptionfrom the samples without added RNA pol II could only come from RNA pol IIpresent in the Mediator preparations.

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The kinase module is believed to function as an inhibitor oftranscription while Mediator associated with RNA pol II isbelieved to be active. Based on NSAF values, statistical analysis,and quantitative Western blotting, F-Med26 and F-Med10 Me-diator had enriched levels of RNA pol II, whereas F-Med26Mediator preparations were depleted of kinase module com-pared with F-Med10 (Fig. 2). F-Med28 and F-Med29 Mediatorhad similar levels of kinase module compared with F-Med10, butdiminished levels of RNA pol II components. These resultspredict that F-Med26 Mediator should have increased activitywhen compared with the other preparations. As an independentmeasurement of functional RNA pol II present in our prepara-tions of Mediator we performed in vitro transcription assaysusing a highly purified, reconstituted transcription system. Un-der the reaction conditions used in this assay, none of theMediator preparations affected basal transcription from theAdenovirus 2 major late promoter when reactions containedsaturating amounts of TBP, TFIIB, TFIIE, TFIIF, TFIIH, andpurified pol II (Fig. 3C, lanes 26). When purified pol II wasomitted, however, transcription was highest in the presence ofMediator purified through F-Med26 followed by F-Med10 (Fig.3C, lanes 7 and 8), consistent with NSAF data indicating thatthese preparations contained the highest amounts of RNA pol II,while very little transcription was observed in reactions withF-Med28 or F-Med29 Mediator. A possible explanation for thedifference in activity between F-Med26 and F-Med10 Mediatorsis the F-Med26 population contains slightly more RNA polcomponents coupled with dramatically less kinase module com-pared with the F-Med10 preparations, as supported by the kinasemodule andRNA pol II NSAF analysis and quantitativeWesternblotting. Furthermore, of the five RNA pol II componentsstatistically different between F-Med26 and F-Med10 prepara-tions, four were increased in abundance in F-Med26 Mediator(Rpb1, 3, 7, and 9), while only Rpb5 was increased in abundancein F-Med10 preparations (Fig. 3 and SI).

DiscussionIn this report we have demonstrated a statistically robust ap-proach for structure function analyses of multiprotein complexesusing label-free quantitativeMudPIT andmammalianMediator.Conclusions drawn from our quantitative analyses of MudPITdata using NSAFs were validated by an AQUA experiment (36)

using 15N-labeled Med9, quantitative Western blotting, andtranscription assays. Hence, with the use of NSAF, MudPIT canbe a powerful tool not only for protein identification but also forobtaining information about the abundance of proteins presentin purified multiprotein complexes. The most striking aspect ofthe data is that the subunit that is FLAG-tagged has a sizableimpact on the makeup of bulk Mediator that is purified. Basedon the results of our NSAF analyses we conclude that there islittle significant difference in the relative stoichiometry of mostMediator subunit complexes purified through any of four dif-ferent baits, Med26, Med10, Med28, and Med29. These findingsdefine a common core of Mediator subunits that are likelypresent in all or most Mediator complexes found in cells.We nevertheless find significant differences in the apparent

compositions of complexes purified through different epitope-tagged subunits, arguing for the existence of multiple forms ofMediator complex that share a common core of subunits. Basedon the MS data from this study, Mediator purified through theseFLAG-tagged subunits falls into three main complexes (Fig. 4).On one end of the spectrum is the F-Med26 Mediator, whichresembles the previously described cofactor required for Sp1(57, 19, 20, 26) and the positive cofactor 2 (17, 18) purificationsthat have been reported to be highly active in support ofactivated transcription. This complex appears to be almostcompletely lacking in kinase module, with a high percentage ofassociated RNA pol II (Fig. 4A). It is important to note that asmall amount of kinase module subunits were isolated in asso-ciation with F-Med26; hence, although the majority of Mediatorthat contain Med26 lack kinase module, it is possible for bothMed26 and the kinase module to interact simultaneously withthe core complex.On the other end of the spectrum are the F-Med28/Med29

complexes that resemble the thyroid hormone receptor-associated protein complex (3, 14, 21, 22), SRB/Med-containingcofactor complex (9), negative regulator of activated transcrip-tion (16), vitamin D receptor interacting protein complex (12,13), SRB (8, 15), and activator recruited cofactor-L (10, 11)purifications described previously and likely represent the ma-jority of the transcriptionally repressed Mediator present in thecells. Based on NSAF analyses of MudPIT data and assays forRNA pol II activity we found that Mediator purified throughF-Med28 and F-Med29 is apparently associated with less RNA

Fig. 4. Bait-dependent distribution of Mediator, kinase, and RNA pol modules. (A) The NSAF values for each protein in each of the three overall modules(Mediator, kinase, and RNA pol II) was summed as percentage of whole determined for each Mediator bait used. Filled squares, open circles, and filled circlesrepresent the percent of the complex that was Mediator, RNA pol II, and kinase module, respectively. (B) A proposed model for the states of Mediator in a cellularenvironment where Mediator with the kinase module is transcriptionally inactive and Mediator with the RNA pol II module is transcriptionally active. Animportant question is whether the pol and kinase modules can interact with the same Mediator molecule or are mutually exclusive.

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pol II than Mediator purified through F-Med10 (Fig. 4A).Association of F-Med28 and F-Med29 with Mediator clearlydoes not prevent binding of RNA pol II (or vice versa), sinceMed28 and Med29 do not appear to be depleted in Mediatorpurified through F-Med26, which is associated with a largeamount of RNA pol II. Association of the kinase module withMediator has been proposed to block binding of RNA pol II;however, Mediator purified through F-Med28 or F-Med29 doesnot contain substantially more kinase module than Mediatorpurified through F-Med10.These F-Med28/Med29 Mediator complexes incorporate a

relatively high percentage of kinase module, with little associ-ated RNA pol II (Fig. 4A). The F-Med10 preparation appearsto offer a multifaceted picture, both in terms of complexcomposition and functionality. It is possible that this preparationeither is composed of (i) one large complex incorporatingMediator with kinase module and RNA pol II or (ii) severalsmaller complexes, Mediator/pol II, Mediator/kinase moduleand Mediator alone as well as the larger complex describedabove (Fig. 4B). Diverse functionality can be hypothesized forthe larger complex, which is not mutually exclusive and cannotbe ruled out by the present literature. The larger complex(Mediator/kinase/pol II) can potentially represent both an activeand repressive form of Mediator, governed by small changes insubunit composition that are promoter specific (21), or changesin overall three dimensional structure when bound to differentactivators (26). Another possibility is that this larger complexrepresents a transition from active to repressive Mediator (orvice versa). There is support in the literature for repressedMediator, after stimuli from an activator, to lose the kinasemodule, incorporate Med26 and bind RNA pol II in a stepwisefashion, before transcription initiation (23). It is possible that atransition state exists that incorporates all of these proteins (4).Further fractionation of the F-Med10 preparation (along withMudPIT analysis and transcription assays) has the potential toclearly delineate overall Mediator composition and the func-tionality associated to each complex.From an analytical standpoint, the method of protein complex

characterization described in this article has important implica-tions. MudPIT is routinely used to identify proteins in complexes(27), but we demonstrate a statistically robust approach forquantitative analysis of multiprotein complexes that yields struc-ture/function insights. By replicate analysis of multiprotein Me-diator complexes using distinct Mediator proteins as baits toanalyze the complex, the NSAF approach revealed uniqueprotein abundances and complex structures. Available antibod-ies validated these results and the functional significance wasdemonstrated with a transcription assay. The NSAF approachfor relative abundance determination has excellent potential forstatistically robust structure function analyses of multiproteincomplexes where existing methods are unable to provide com-prehensive information and a full suite of antibodies is notavailable.

Materials and MethodsRecombinant 15N Med9 Purification. Recombinant Med9 clonedinto a pET vector in E. coli strain BL21(DE3) (Novagen,Madison, WI) and expressed as a 6His fusion protein (Nova-gen) was grown in Bioexpress cell growth media (U-15N, 98%)(Cambridge Isotope Laboratories, Andover,MA). Recombinant15N Med9 was purified from E. coli extracts first by Ni-NTAagarose chromatography (Qiagen, Valencia, CA), then by ionexchange chromatography on a TSKgel DEAE-5PW column(Tosoh Biosciences, Tokyo, Japan) using a 25-min 0.0050.25 Msodium chloride gradient generated by a Beckman CoulterSystem Gold HPLC (Beckman Coulter, Fullerton, CA). Proteinconcentration was determined by protein assay (Bio-Rad, Her-cules, CA), and 5 g of purified 15N Med9 was trypsin-digested

(Roche, Mannheim, Germany) and analyzed on a LCQDeca XPPlus mass spectrometer (Thermo Electron, Waltham, MA) todetermine purity based on percentage of Med9 spectral counts(87%).

Mediator Preparations and MudPIT Analysis. Nuclear extracts wereprepared according to the method of Dignam et al. (41) from 6109 HeLa cells stably expressing N-terminally FLAG-taggedMed26 (formerly Crsp70), Med29 (formerly Intersex), Med28,or Med10 (formerly Nut2) subunits. Nuclear extracts weresubjected to anti-FLAG agarose immunoaffinity chromatogra-phy (Sigma, St. Louis, MO) as described in ref. 27 and elutedwith FLAG peptide (Sigma). Four independent preparations ofMediator complex were generated from each FLAG-tagged cellline. Recombinant 15N Med9 was added to all four replicates ofall four FLAG-tagged preparations in various amounts as fol-lows: 0.113 g added to F-Med26, 0.128 g added to F-Med29,0.018 g added to F-Med28, and 0.042 g added to F-Med10Mediator.Each sample was digested with trypsin (Roche) and analyzed

byMudPIT as described previously (28, 42) on an LTQ linear iontrap mass spectrometer (Thermo Electron, San Jose, CA).Variations to standardMudPIT approaches wheremixtures wereresolved by an eight-step chromatographic program consisting of5% buffer C (500 mM ammonium acetate, 5% acetonitrile, and0.1% formic acid), 15% buffer C, 30% buffer C, 50% buffer C,50% buffer C, 70% buffer C, and two successive steps using100% buffer C as the salt bumps within standard MudPITchromatographic programs (42).Data sets were searched by using SEQUEST against a data-

base of 56,838 protein sequences, combining 28,242 Homosapiens proteins (from the National Center for BiotechnologyInformation, February 17, 2005), 177 common contaminants likekeratin and Igs, and their corresponding shuffled sequences (i.e.,each H. sapiens and contaminant sequence was randomizedmaintaining the same amino acid composition and length).SEQUEST was run to search for methionine oxidation and withno enzyme specificity, an approach that has been demonstratedto be superior for minimizing false positive identifications (43).Tandem MS data sets were searched twice to match spectra topeptides generated from endogenous 14N Mediator protein andto 15N peptides derived from the spiked recombinant Med19protein (29, 35). Protein lists were generated by using DTASelect(44) based on the following filtering criteria: each peptide had tobe at least 7 aa long and fully tryptic, with a Cn of at least 0.08,and minimum Xcorr of 1.5 for 1, 1.9 for 2, and 2.6 for 3charged precursor ions (45). Ambiguous peptides and redundantproteins were removed by using DTASelect (44). By using theparameters described in the methods, the average peptide falsepositive rates according to the equation used by Elias et al. (45)were 0.85% for Med10, 0.80% for Med26, 0.5% for Med28, and1.3% for Med29. The peptide identifications from a represen-tative run of each analysis are provided in the SI.Spectral count normalization was applied to proteins belonging

to the Mediator or RNA pol II complexes to estimate the relativelevels of each subunit within the complexes. Here, for each proteink involved in Mediator/pol II, we calculated a SAF normalizedagainst the whole protein complex (NSAF) as follows:

(NSAF)k SpCLk

i1N SpCL i

,

in which the total number of tandem MS spectra matchingpeptides from protein k (SpC) was divided by the proteins length(L), then divided by the sum of SpC/L for all N proteins inMediator (32 subunits) and pol II (12 subunits).

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Western Blot Analyses. To compare levels of individual subunitspresent in each sample, Western blot analyses were performed andnormalized to the level of Med17, because MS data demonstratedthat this subunit should be of similar abundance in each prepara-tion. Once the amount of individual Mediator preparation neededto produce equal blotting of Med17 was determined, all Westernblot analyses were produced by using the same volumes. Proteinswere transferred onto poly(vinylidene difluoride) and blotted byusing primary antibodies against Med17 and Med18 (CocalicoBiologicals, Reamstown, PA), Med8 (46), Cdk8 (Santa Cruz Bio-technology, Santa Cruz, CA), cyclin C (Neomarkers, Fremont,CA), Rpb1 and Rpb9 (Protein One, BethesdaMD), and secondaryantibodies against mouse or rabbit IgG (GE Healthcare, Piscat-away, NJ). Blots were developed with the ECL Plus WesternBlotting Detection System (GEHealthcare) and imaged by using aTyphoon 9400 (GE Healthcare).

Transcription Assays. Transcription reactions were performed asdescribed (37) with the EcoR1 to NdeI fragment of pDN-AdML

as template purified RNA pol II from calf thymus (a kind gift ofAvi Gnatt, University of Maryland School of Medicine, Balti-more, MD) and TFIIH from rat liver, and recombinant TBP,TFIIB, TFIIE, and TFIIF. Template, RNA pol II, and transcrip-tion factors were incubated at 28C for 30 min to allow forma-tion, and transcription was initiated by addition of 50 M ATP,UTP, and GTP, 6 M CTP, 7 Ci of [-32P]CTP (600 Ci/mmol;GE HealthCare), and 8 mM MgCl2. After 30 min at 28C,reactions were stopped by addition of EDTA to 0.25 mM, NaClto 0.15 M, 0.02 mg of Proteinase K, and 0.04 mg of tRNA.Reaction products were subjected to electrophoresis on a 6%polyacrylamide (19:1 acrylamide:bisacrylamide) gel containing 6M urea and 0.5 TBE and imaged with Typhoon 8600 (Molec-ular Dynamics).

This work was supported by the Stowers Institute for MedicalResearch and by National Institutes of Health Grant R37GM41628(to R.C.C.).

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Paoletti et al. PNAS December 12, 2006 vol. 103 no. 50 18933

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Table 2. Representative MudPIT analysis results for Med10(Nut2), Med26(Crsp70), Med28, and Med29(Intersex) Mediator and RNA Polymerase II components

DTASelect v1.9 Parameters Used for All Runs Locations Mediator Nomenclature Key1.5 Minimum +1 XCorr Med10(Nut2) starts on line 41 Consensus In Files1.9 Minimum +2 XCorr Med26(Crsp70) starts on line 1419 Cdk8L Cdk112.6 Minimum +3 XCorr Med28 starts on line 2249 Cdk8 Cdk80.08 Minimum DeltCN Med29(Intersex) starts on line 3009 cyclin C cyclin C

1 Minimum charge state Med1 TRAP2203 Maximum charge state Med10 Nut20 Minimum ion proportion Med11 HSPC296

10 Maximum Sp rank Med12 TRAP230-1 Minimum Sp score Med13 TRAP240S

Include Modified peptide inclusion Med13L TRAP240LFull Tryptic status requirement Med14 Rgr1

FALSE Multiple, ambiguous IDs allowed Med15 ARC1057 Minimum sequence length Med16 TRAP95

100 Maximum sequence length Med17 TRAP80XCorr Purge duplicate peptides by protein Med18 p28b

FALSE Include only loci with unique peptide Med19 LCMR1TRUE Remove subset proteins Med20 TRFP

Contaminant Exclude protein names matching Med21 SRB710 Minimum redundancy for low coverage loci Med22 Surf51 Minimum peptides per locus Med23 Sur2

Med24 TRAP100Med25 ARC92Med26 CRSP70Med27 TRAP37Med28 AK007855Med29 IntersexMed30 TRAP25Med31 SOH1Med4 TRAP36Med6 Med6Med7 Med7Med8 Med8Med9 Med25

Med10(Nut2) Representative Analysis of Mediator and RNA Polymerase II ComponentsLocus Sequence Count Spectrum CSequence Length MolWt pI Validation SDescriptive NameUnique FileName XCorr DeltCN M+H+ CalcM+H+ TotalIntensitySpRank SpScore IonProportiRedundancSequencegi|21361161|re 11 248 86.10% 144 15564 4.4 U* HsNut2Med_Ti_03.000039.000039.3 7.9701 0.5358 4361.15 4361.788 6465.9 1 2254.2 33.3 42 R.LTQLQDAVNSLADQFCNAIGVLQQCGPPASFNNIQTAINK.D* HsNut2Med_Ti_02.017120.017120.2 6.7037 0.4351 2219.37 2219.459 4916.2 1 1587.4 76.3 78 K.DQPANPTEEYAQLFAALIAR.T* HsNut2Med_Ti_02.016562.016562.3 6.3864 0.3853 2220.2 2219.459 6460.3 1 1657.6 46.1 26 K.DQPANPTEEYAQLFAALIAR.T* HsNut2Med_Ti_05.000231.000231.2 6.7602 0.5313 2650.21 2650.94 4440.5 1 1095.9 58.3 28 K.DIDVLIDSLPSEESTAALQAASLYK.L* HsNut2Med_Ti_02.014894.014894.3 4.9653 0.446 2650.76 2650.94 5648.6 1 931 34.4 4 K.DIDVLIDSLPSEESTAALQAASLYK.L* HsNut2Med_Ti_08.009763.009763.2 5.4982 0.4805 2177.19 2178.283 9674.4 1 2041.1 67.6 18 K.LEEENHEAATCLEDVVYR.G* HsNut2Med_Ti_04.007760.007760.3 4.7511 0.2919 2178.11 2178.283 9053.8 2 1860.3 44.1 9 K.LEEENHEAATCLEDVVYR.G* HsNut2Med_Ti_05.002675.002675.3 4.9117 0.3152 2979.32 2981.225 5167.9 1 1197.8 39.6 6 K.LEEENHEAATCLEDVVYRGDM@LLEK.I* HsNut2Med_Ti_04.009303.009303.2 3.8403 0.3842 2979.51 2981.225 4486.4 1 284.1 41.7 2 K.LEEENHEAATCLEDVVYRGDM@LLEK.I* HsNut2Med_Ti_03.008495.008495.1 3.2987 0.3211 1486.54 1486.709 5564.9 1 734.7 65.4 3 K.IQSALADIAQSQLK.T* HsNut2Med_Ti_02.018429.018429.2 5.0776 0.3401 1486.81 1486.709 8391.7 1 2257.4 84.6 32 K.IQSALADIAQSQLK.Tgi|29840782|sp 23 112 77.60% 246 28425 8.6 Ugi|42544155|re 23 112 77.60% 246 28425 8.6 U MEDIATOR_MED6 {mediator of RNA polymerase II transcription, subunit 6 homolog} [Homo sapiens]

HsNut2Med_Ti_07.015665.015665.3 3.3566 0.0845 3282.86 3284.649 10706.8 5 519.3 21.4 1 R.DNLLGISWVDSSWIPILNSGSVLDYFSER.SHsNut2Med_Ti_04.006717.006717.2 3.4868 0.3274 1841.25 1843.957 9331.3 2 749.4 53.6 1 R.SNPFYDRTCNNEVVK.MHsNut2Med_Ti_08.013586.013586.3 6.9308 0.3879 3336.86 3334.989 7486.7 1 1688.3 38 2 R.LTLEHLNQM@VGIEYILLHAQEPILFIIR.KHsNut2Med_Ti_03.018254.018254.3 8.3584 0.3896 3435.86 3433.929 5994.2 1 1350.2 34.7 38 R.QSPAQVIPLADYYIIAGVIYQAPDLGSVINSR.VHsNut2Med_Ti_05.012114.012114.2 5.2347 0.5451 2255.49 2256.504 9281.3 1 2394.3 71.1 3 R.VLTAVHGIQSAFDEAMSYCR.YHsNut2Med_Ti_05.012105.012105.3 3.5941 0.2729 2255.6 2256.504 8496.9 1 1050.8 34.2 2 R.VLTAVHGIQSAFDEAMSYCR.YHsNut2Med_Ti_05.010052.010052.2 5.5695 0.5696 2271.33 2272.504 6377.7 1 1369.2 65.8 9 R.VLTAVHGIQSAFDEAM@SYCR.YHsNut2Med_Ti_04.008757.008757.3 5.265 0.3324 2271.65 2272.504 7521.9 1 2064.8 46.1 16 R.VLTAVHGIQSAFDEAM@SYCR.YHsNut2Med_Ti_07.009188.009188.3 4.2248 0.2147 1378.22 1377.543 4933.1 1 905.5 62.5 2 K.RKEEPSSIFQR.QHsNut2Med_Ti_07.009182.009182.2 3.6196 0.1074 1378.49 1377.543 4944.4 2 698.1 80 1 K.RKEEPSSIFQR.QHsNut2Med_Ti_04.006483.006483.2 2.696 0.1095 1222.07 1221.356 5882 1 776.5 83.3 2 R.KEEPSSIFQR.QHsNut2Med_Ti_05.009854.009854.2 2.9125 0.1382 1314.17 1312.555 3498.5 4 554 75 7 R.QRVDALLLDLR.QHsNut2Med_Ti_02.009998.009998.2 3.5146 0.1862 1028.27 1028.237 6414.9 4 1328.6 87.5 4 R.VDALLLDLR.QHsNut2Med_Ti_06.007905.007905.2 2.5481 0.1301 1910.19 1911.252 7336.7 2 475.1 50 1 K.FVQLKPGEKPVPVDQTK.KHsNut2Med_Ti_08.008849.008849.3 5.1154 0.2774 2040.29 2039.426 7965.1 1 2116.2 48.5 6 K.FVQLKPGEKPVPVDQTKK.EHsNut2Med_Ti_08.008937.008937.2 4.549 0.193 2041.47 2039.426 6056.2 1 852.6 58.8 2 K.FVQLKPGEKPVPVDQTKK.EHsNut2Med_Ti_04.005044.005044.2 3.6616 0.24 1293.63 1295.478 4250.3 1 790.4 72.7 1 K.PGEKPVPVDQTK.KHsNut2Med_Ti_03.006160.006160.2 3.3822 0.2258 1724.07 1724.949 6420.6 6 468.2 53.6 2 K.KEAEPIPETVKPEEK.EHsNut2Med_Ti_03.005854.005854.3 4.0455 0.3488 1724.24 1724.949 7448.4 1 989.8 44.6 2 K.KEAEPIPETVKPEEK.EHsNut2Med_Ti_02.005864.005864.1 3.1786 0.3488 1595.45 1596.775 2681 1 297.2 65.4 2 K.EAEPIPETVKPEEK.EHsNut2Med_Ti_02.006116.006116.2 3.4502 0.184 1597.17 1596.775 3733.9 1 623.2 76.9 4 K.EAEPIPETVKPEEK.EHsNut2Med_Ti_03.005830.005830.2 4.4603 0.4348 2056.15 2056.274 4864.7 1 533.8 55.9 2 K.EAEPIPETVKPEEKETTK.NHsNut2Med_Ti_02.003644.003644.2 2.4668 0.1802 974.55 975.09 4531.3 2 784.1 81.2 2 K.NVQQTVSAK.G

gi|19557695|re 10 60 74.50% 200 22221 4.7 U surfeit 5 isoform b; surfeit locus protein 5 [Homo sapiens] HsNut2Med_Ti_02.006368.006368.1 2.8833 0.1428 1096.36 1096.225 3347.5 1 252.3 75 2 K.ETLLQSYNK.RHsNut2Med_Ti_02.011376.011376.2 2.8748 0.0854 1311.09 1311.536 6714.1 5 617.4 70 1 K.SIMDNFTEIIK.THsNut2Med_Ti_02.008728.008728.2 3.6499 0.2666 1325.81 1327.536 5468.9 2 876.8 85 3 K.SIM@DNFTEIIK.THsNut2Med_Ti_03.004902.004902.1 2.7726 0.2372 1377.41 1377.494 5335 1 320.5 54.5 2 K.TAKIEDETQVSR.AHsNut2Med_Ti_03.004703.004703.2 3.6087 0.1999 1380.19 1377.494 7660.8 1 1461.5 81.8 2 K.TAKIEDETQVSR.AHsNut2Med_Ti_03.004100.004100.2 4.1375 0.3922 1694.19 1694.774 5681.4 1 765.6 69.2 4 R.ATQGEQDNYEM@HVR.AHsNut2Med_Ti_03.013382.013382.2 5.8884 0.3716 2120.25 2120.37 6368.8 1 1700.7 73.5 38 K.QFLILNDFPSVNEAIDQR.N

MEDIATOR_SBR7 {SRB7 suppressor of RNA polymerase B homolog; RNA polymerase II complex component SRB7; SRB7 (suppressor of RNA polymerase B, yeast) homolog} [Homo sapiens]

MEDIATOR_MED6 {MED6_HUMAN RNA polymerase transcriptional regulation mediator, subunit 6 homolog (Activator-recruited cofactor 33 kDa component) (ARC33) (NY-REN-28 antigen)}

HsNut2Med_Ti_03.013283.013283.3 5.8837 0.2796 2121.53 2120.37 9232.4 1 3866 57.4 6 K.QFLILNDFPSVNEAIDQR.N* HsNut2Med_Ti_08.014043.014043.3 3.6805 0.1493 4718.72 4719.053 7419.1 1 420.1 18.6 1 K.LITLRDEISIDLYELEEEYYSSSSSLCEANDLPLCEAYGR.L* HsNut2Med_Ti_06.010948.010948.3 3.6701 0.2106 4296.29 4297.638 8655.7 1 655.2 19.3 1 R.LDLDTDSADGLSAPLLASPEPSAGPLQVAAPAHSHAGGPGPTEHA.-gi|10863925|re 4 18 73.10% 67 7645 7.8 U PolII_Rpb10 {DNA directed RNA polymerase II polypeptide L; polymerase (RNA) II (DNA directed) polypeptide L (7.6kD); RNA polymerase II subunit} [Homo sapiens] * HsNut2Med_Ti_07.013526.013526.3 5.6195 0.413 3150.47 3152.571 9451.5 1 2016.1 33.9 5 K.IVGNKWEAYLGLLQAEYTEGDALDALGLK.R* HsNut2Med_Ti_03.016707.016707.2 5.262 0.47 2639.43 2640.949 11628.1 1 2050.5 52.2 3 K.WEAYLGLLQAEYTEGDALDALGLK.R* HsNut2Med_Ti_08.009779.009779.2 4.229 0.124 1301.29 1298.584 5587.4 1 1469.6 90 9 R.M@LLAHVDLIEK.L* HsNut2Med_Ti_03.007280.007280.2 2.812 0.1682 1063.41 1061.266 3608.9 4 720.8 87.5 1 K.LLNYAPLEK.-gi|38258656|sp 9 108 72.50% 131 15805 8.5 U MEDIATOR_SOH1 {SOH1_HUMAN Mediator complex subunit SOH1 (hSOH1) (CGI-125)} gi|7705592|ref|NP_057144.1| [Homo sapiens] gi|7705592|ref| 9 108 72.50% 131 15805 8.5 U MEDIATOR_SOH1 {CGI-125 protein} [Homo sapiens]

HsNut2Med_Ti_06.017798.017798.2 4.8871 0.3885 2826.79 2829.194 4599.6 1 824.4 52.3 34 R.FQLELEFVQCLANPNYLNFLAQR.GHsNut2Med_Ti_07.014025.014025.3 5.8782 0.359 2828.09 2829.194 11613.7 1 2162.9 37.5 18 R.FQLELEFVQCLANPNYLNFLAQR.GHsNut2Med_Ti_05.009135.009135.2 3.1378 0.123 1591.21 1589.831 7086.7 1 847 72.7 1 K.YLLYWKDPEYAK.YHsNut2Med_Ti_08.010587.010587.2 4.7603 0.4194 2294.81 2294.6 3748.9 1 581.3 68.8 3 K.YPQCLHM@LELLQYEHFR.KHsNut2Med_Ti_07.010922.010922.3 4.6362 0.2591 2294.84 2294.6 5989.6 1 1152.3 48.4 2 K.YPQCLHM@LELLQYEHFR.KHsNut2Med_Ti_02.004221.004221.1 2.5327 0.0952 1034.3 1033.136 2596.2 1 254 75 1 K.ELVNAQCAK.FHsNut2Med_Ti_08.010265.010265.2 4.6714 0.3356 2002.29 2001.211 7550.1 1 1632 71.4 7 K.FIDEQQILHWQHYSR.KHsNut2Med_Ti_07.008789.008789.2 5.0255 0.431 2188.53 2190.396 3818.7 1 623.9 63.9 3 R.M@RLQQALAEQQQQNNTSGK.-HsNut2Med_Ti_03.004749.004749.2 6.145 0.4382 1887.23 1887.016 8303.2 1 1923.5 71.9 39 R.LQQALAEQQQQNNTSGK.-

gi|14702171|re 15 65 71.30% 275 31441 4.9 U PolII_Rpb3 {DNA directed RNA polymerase II polypeptide C; polymerase (RNA) II (DNA directed) polypeptide C (33kD); RNA polymerase II subunit 3} [Homo sapiens] gi|14702173|re 15 65 71.30% 275 31441 4.9 U PolII_Rpb3 {DNA directed RNA polymerase II polypeptide C; polymerase (RNA) II (DNA directed) polypeptide C (33kD); RNA polymerase II subunit 3} [Homo sapiens]

HsNut2Med_Ti_02.005751.005751.1 2.8811 0.179 1161.44 1162.282 6098.3 1 677 77.8 2 R.ITELTDENVK.FHsNut2Med_Ti_02.010022.010022.2 5.3353 0.3488 1675.55 1676.91 7663.4 1 2020.5 82.1 16 K.FIIENTDLAVANSIR.RHsNut2Med_Ti_08.013934.013934.3 5.3003 0.364 3520.28 3520.025 3086.8 1 376 28.3 5 R.VFIAEVPIIAIDWVQIDANSSVLHDEFIAHR.LHsNut2Med_Ti_02.013096.013096.1 3.1575 0.2976 1511.58 1511.799 6487.6 1 1010.1 69.2 4 R.LGLIPLISDDIVDK.LHsNut2Med_Ti_03.013254.013254.2 4.23 0.3258 1512.55 1511.799 2876.7 1 765.7 84.6 7 R.LGLIPLISDDIVDK.LHsNut2Med_Ti_03.012359.012359.2 2.3611 0.1257 2553.73 2554.588 5889.1 1 177.1 36.8 2 R.DCTCEEFCPECSVEFTLDVR.CHsNut2Med_Ti_03.008818.008818.2 5.4235 0.4047 2276.05 2276.422 5450.5 1 1392.3 69.4 9 R.NRDNDPNDYVEQDDILIVK.LHsNut2Med_Ti_06.008900.008900.3 4.6515 0.1581 2278.07 2276.422 7868.7 1 2157.2 48.6 2 R.NRDNDPNDYVEQDDILIVK.LHsNut2Med_Ti_02.010725.010725.2 5.0647 0.508 2039.33 2037.196 6780.5 1 1014.9 61.8 3 K.WNPTAGVAFEYDPDNALR.HHsNut2Med_Ti_02.007187.007187.2 3.5003 0.1905 2656.19 2656.689 5582.3 1 478 43.2 1 K.SEYSELDEDESQAPYDPNGKPER.FHsNut2Med_Ti_02.006692.006692.3 4.3573 0.3598 2657.69 2656.689 7172.5 1 1043.2 34.1 2 K.SEYSELDEDESQAPYDPNGKPER.FHsNut2Med_Ti_06.015791.015791.3 4.3125 0.326 2803.76 2805.166 8300.2 1 2295.7 40.6 5 R.FYYNVESCGSLRPETIVLSALSGLK.KHsNut2Med_Ti_04.009764.009764.3 2.6985 0.1599 2381.69 2383.661 4560.9 1 741.8 37.5 1 K.KLSDLQTQLSHEIQSDVLTIN.-HsNut2Med_Ti_04.009734.009734.2 4.9406 0.3377 2384.03 2383.661 5018.2 1 455.6 50 5 K.KLSDLQTQLSHEIQSDVLTIN.-HsNut2Med_Ti_03.012668.012668.2 3.8213 0.2688 2254.79 2255.487 6220.5 2 350.3 42.1 1 K.LSDLQTQLSHEIQSDVLTIN.-

gi|7661788|ref| 28 228 68.90% 270 29745 5.1 U MEDIATOR_TRAP36 {vitamin D receptor interacting protein; HSPC126 protein; p36 TRAP/SMCC/PC2 subunit} [Homo sapiens] * HsNut2Med_Ti_06.006482.006482.2 4.9487 0.3544 1500.75 1501.643 3227.6 1 798.7 80 2 K.ERLGGGLGVAGGNSTR.E* HsNut2Med_Ti_06.006473.006473.3 3.6177 0.2295 1501.19 1501.643 5410.5 1 1172 45 2 K.ERLGGGLGVAGGNSTR.E* HsNut2Med_Ti_04.006086.006086.1 3.2926 0.2801 1215.47 1216.34 2735.1 1 261.4 61.5 4 R.LGGGLGVAGGNSTR.E* HsNut2Med_Ti_04.006170.006170.2 4.2457 0.3703 1217.25 1216.34 5369.7 1 1627.2 88.5 4 R.LGGGLGVAGGNSTR.E* HsNut2Med_Ti_06.013793.013793.2 4.182 0.3377 1742.47 1743.998 4614 1 836.8 75 11 R.ERLLSALEDLEVLSR.E* HsNut2Med_Ti_07.012365.012365.3 4.8351 0.1306 1744.82 1743.998 6000.7 6 1162.5 46.4 1 R.ERLLSALEDLEVLSR.E* HsNut2Med_Ti_02.012812.012812.2 4.9055 0.3707 1458.49 1458.695 9765.6 1 2633.5 87.5 46 R.LLSALEDLEVLSR.E* HsNut2Med_Ti_03.013005.013005.3 4.6259 0.2056 1458.53 1458.695 8729.2 4 2514.7 62.5 2 R.LLSALEDLEVLSR.E* HsNut2Med_Ti_02.012849.012849.1 3.0671 0.1867 1459.7 1458.695 4561 1 613.1 70.8 9 R.LLSALEDLEVLSR.E* HsNut2Med_Ti_05.008678.008678.2 3.3394 0.1188 1192.21 1191.429 7865.3 3 1364 88.9 13 R.ELIEM@LAISR.N* HsNut2Med_Ti_06.012728.012728.2 5.6296 0.3582 1974.19 1976.283 9077.4 1 2560.9 75 23 K.LLQAGEENQVLELLIHR.D* HsNut2Med_Ti_07.011972.011972.3 4.195 0.3918 1974.68 1976.283 5648.1 1 1315.3 51.6 18 K.LLQAGEENQVLELLIHR.D* HsNut2Med_Ti_07.012004.012004.3 4.4959 0.308 3071.3 3070.488 5933.4 1 829.6 33 4 K.LLQAGEENQVLELLIHRDGEFQELM@K.L* HsNut2Med_Ti_02.007750.007750.1 2.1243 0.152 1096.46 1097.228 6054.1 9 479.9 68.8 2 R.DGEFQELMK.L* HsNut2Med_Ti_02.007731.007731.2 2.9779 0.2047 1097.17 1097.228 5907.7 5 1045.8 87.5 1 R.DGEFQELMK.L* HsNut2Med_Ti_02.006278.006278.1 2.1653 0.0951 1112.37 1113.228 2407.5 1 157.8 68.8 2 R.DGEFQELM@K.L* HsNut2Med_Ti_01.008862.008862.2 2.6021 0.1087 1113.55 1113.228 4997.1 1 619.6 87.5 1 R.DGEFQELM@K.L* HsNut2Med_Ti_02.008036.008036.1 3.8223 0.456 1534.47 1535.737 2955.1 1 320.3 65.4 7 K.EAEQILATAVYQAK.E* HsNut2Med_Ti_04.007910.007910.2 5.1589 0.3568 1537.01 1535.737 10512 1 2368.6 76.9 41 K.EAEQILATAVYQAK.E* HsNut2Med_Ti_02.006320.006320.2 3.1322 0.1092 1047.11 1047.194 4544.1 4 844.1 77.8 2 K.GAISSEEIIK.Y* HsNut2Med_Ti_03.009662.009662.3 4.4042 0.2697 2010.89 2012.239 4026.2 1 1825.5 52.8 4 R.ISASNAVCAPLTWVPGDPR.R* HsNut2Med_Ti_02.009626.009626.2 4.8471 0.4398 2012.29 2012.239 5743.6 1 1347.9 61.1 6 R.ISASNAVCAPLTWVPGDPR.R* HsNut2Med_Ti_06.006340.006340.1 2.0469 0.2013 1293.31 1294.469 3452.8 1 226.6 66.7 1 R.RPYPTDLEM@R.S* HsNut2Med_Ti_06.006357.006357.3 2.6723 0.1416 1294.4 1294.469 6254.4 2 1066.9 61.1 1 R.RPYPTDLEM@R.S* HsNut2Med_Ti_04.008202.008202.3 3.5406 0.0911 2722.13 2719.992 8549.7 1 874 29.6 1 R.SGLLGQMNNPSTNGVNGHLPGDALAAGR.L* HsNut2Med_Ti_05.008480.008480.3 5.1431 0.254 2734.4 2735.992 6332.3 1 1137.7 34.3 7 R.SGLLGQM@NNPSTNGVNGHLPGDALAAGR.L* HsNut2Med_Ti_07.009898.009898.2 3.8528 0.3953 2734.59 2735.992 5278.6 1 378 37 9 R.SGLLGQM@NNPSTNGVNGHLPGDALAAGR.L* HsNut2Med_Ti_06.011769.011769.3 3.9524 0.2503 4367.57 4367.871 5599.5 2 273.6 17.6 4 R.LPDVLAPQYPWQSNDM@SM@NM@LPPNHSSDFLLEPPGHNK.Egi|28559039|re 110 753 65.30% 1581 168477 8.7 U

HsNut2Med_Ti_07.009406.009406.1 2.5166 0.1092 1519.48 1520.688 3523.4 5 239.4 54.5 1 K.FNQNRPWSETIK.LHsNut2Med_Ti_06.007166.007166.2 3.6423 0.1785 1521.25 1520.688 5390 1 683.5 72.7 4 K.FNQNRPWSETIK.LHsNut2Med_Ti_07.009513.009513.2 4.6297 0.3653 2226.27 2227.507 4268.7 1 598.6 57.9 4 R.VVM@SSGGHQHLVSCLETLQK.AHsNut2Med_Ti_02.005160.005160.2 2.686 0.1607 1109.17 1107.268 3903.4 2 665.4 83.3 3 K.VTSLPAM@TDR.LHsNut2Med_Ti_06.013670.013670.3 4.9619 0.3563 4205.84 4208.53 8679.7 1 720.8 21.6 16 R.QNGLGSHLSASGTECYITSDM@FYVEVQLDPAGQLCDVK.VHsNut2Med_Ti_07.009532.009532.2 5.4119 0.4007 2172.27 2171.387 8722.1 1 1899.7 69.4 3 K.VAHHGENPVSCPELVQQLR.EHsNut2Med_Ti_03.008714.008714.1 2.5524 0.1713 1416.45 1417.605 6178.3 1 660.4 66.7 2 K.GLVNLYNLPGDNK.LHsNut2Med_Ti_03.008698.008698.2 4.5961 0.2448 1418.99 1417.605 5519.2 1 1260.4 83.3 2 K.GLVNLYNLPGDNK.LHsNut2Med_Ti_05.009170.009170.2 4.5507 0.3362 1659.23 1658.938 7053.4 1 1571.9 75 3 K.GLVNLYNLPGDNKLK.THsNut2Med_Ti_02.012668.012668.2 5.6791 0.3108 1640.25 1639.904 7918.3 1 2833.7 88.5 5 K.MYLALQSLEQDLSK.MHsNut2Med_Ti_02.011285.011285.3 4.4004 0.1976 1656.29 1655.904 7438.1 1 2008.3 57.7 2 K.M@YLALQSLEQDLSK.MHsNut2Med_Ti_02.017741.017741.2 3.866 0.159 1657.41 1655.904 9697.4 1 1558.2 76.9 14 K.M@YLALQSLEQDLSK.MHsNut2Med_Ti_07.009452.009452.1 3.5996 0.3518 1312.5 1313.542 3877.5 1 408.6 72.7 4 K.ILHGSVGYLTPR.SHsNut2Med_Ti_08.002675.002675.2 3.7566 0.4331 1313.07 1313.542 5849.9 1 881.8 72.7 4 K.ILHGSVGYLTPR.S

* HsNut2Med_Ti_01.009768.009768.1 2.5797 0.1809 1415.41 1415.54 6957.8 1 543.6 54.5 8 K.YYVSPSDLLDDK.T* HsNut2Med_Ti_01.009806.009806.2 3.7562 0.3312 1416.41 1415.54 5201.6 1 1362.4 90.9 9 K.YYVSPSDLLDDK.T* HsNut2Med_Ti_05.006682.006682.1 3.9002 0.3753 1550.57 1551.743 4795.6 1 316.5 50 3 K.TASPIILHENNVSR.S* HsNut2Med_Ti_06.006734.006734.3 2.6036 0.2198 1551.83 1551.743 3604.2 9 527.7 42.3 1 K.TASPIILHENNVSR.S* HsNut2Med_Ti_05.006666.006666.2 4.2415 0.3561 1553.09 1551.743 6163.5 1 1169.2 76.9 8 K.TASPIILHENNVSR.S* HsNut2Med_Ti_03.002342.002342.2 5.7803 0.4487 1844.51 1845.075 8376 1 1292.8 61.8 90 R.SLGM@NASVTIEGTSAVYK.L

MEDIATOR_TRAP220 {peroxisome proliferator-activated receptor binding protein; PPARG binding protein; thyroid receptor interacting protein 2; vitamin D receptor-interacting protein 230 kD; thyroid

* HsNut2Med_Ti_03.007263.007263.3 3.9856 0.2996 1845.11 1845.075 9957.5 1 1290.3 39.7 1 R.SLGM@NASVTIEGTSAVYK.L* HsNut2Med_Ti_08.009951.009951.2 3.0822 0.1965 1720.99 1719.053 6820.7 1 785.6 60 2 K.LPIAPLIM@GSHPVDNK.W* HsNut2Med_Ti_02.015038.015038.2 5.7584 0.4768 2578.17 2576.874 8130.8 1 913.1 50 15 K.WTPSFSSITSANSVDLPACFFLK.F* HsNut2Med_Ti_04.000016.000016.3 6.4757 0.4533 3517.64 3516.979 7098.3 1 1916.4 37.1 36 K.LQNCTGIPLFETQPTYAPLYELITQFELSK.D

HsNut2Med_Ti_08.010026.010026.2 4.6294 0.4464 1957.31 1958.193 4041.4 1 775.3 73.3 2 R.FYAALPGQQHCYFLNK.D* HsNut2Med_Ti_05.012447.012447.2 3.5298 0.0879 1051.27 1051.361 5564.7 1 958.2 93.8 27 R.VPLILNLIR.H

HsNut2Med_Ti_07.009584.009584.1 4.1959 0.4268 1589.52 1590.786 4972.1 1 725.2 69.2 3 R.HQVAYNTLIGSCVK.RHsNut2Med_Ti_07.009572.009572.2 4.6293 0.4047 1589.73 1590.786 6544.6 1 1971.7 88.5 4 R.HQVAYNTLIGSCVK.RHsNut2Med_Ti_03.010894.010894.2 5.17 0.4995 2518.43 2519.821 6572.2 1 1153.6 54.8 2 R.TILKEDSPGLLQFEVCPLSESR.FHsNut2Med_Ti_03.010928.010928.3 6.628 0.3279 2520.08 2519.821 9407.2 1 2696.3 45.2 4 R.TILKEDSPGLLQFEVCPLSESR.FHsNut2Med_Ti_01.011171.011171.3 3.4563 0.1975 2063.69 2064.223 6513.7 9 545.1 36.8 1 K.EDSPGLLQFEVCPLSESR.FHsNut2Med_Ti_02.010832.010832.2 5.4537 0.4465 2064.87 2064.223 8265 1 1150.6 58.8 19 K.EDSPGLLQFEVCPLSESR.FHsNut2Med_Ti_08.011713.011713.3 4.5606 0.3348 3326 3323.634 6188.2 1 534.3 27.7 1 R.FSVSFQHPVNDSLVCVVMDVQDSTHVSCK.LHsNut2Med_Ti_07.011698.011698.3 6.2982 0.5011 3337.64 3339.634 5811.9 1 1068.8 33.9 12 R.FSVSFQHPVNDSLVCVVM@DVQDSTHVSCK.LHsNut2Med_Ti_02.010658.010658.2 4.6078 0.3891 1638.47 1639.809 5262.1 1 1219 75 16 K.GLSDALICTDDFIAK.VHsNut2Med_Ti_04.005630.005630.2 2.8052 0.1917 1127.09 1127.327 3772.3 1 501.4 81.2 3 R.CM@SIPVTM@R.AHsNut2Med_Ti_06.013916.013916.3 6.3359 0.4743 2747.96 2747.134 9468.1 1 1860.2 36.5 19 R.RKAETIQADTPALSLIAETVEDM@VK.KHsNut2Med_Ti_07.012290.012290.3 6.2661 0.3783 2875.61 2875.308 6175.9 1 805 35 2 R.RKAETIQADTPALSLIAETVEDM@VKK.NHsNut2Med_Ti_07.014088.014088.2 5.0711 0.392 2577.15 2574.947 9916.1 1 1216.7 47.8 2 R.KAETIQADTPALSLIAETVEDMVK.KHsNut2Med_Ti_06.014096.014096.3 4.8835 0.3367 2590.97 2590.947 6379.5 1 736.7 30.4 6 R.KAETIQADTPALSLIAETVEDM@VK.KHsNut2Med_Ti_03.013599.013599.2 6.4612 0.4924 2592.45 2590.947 5352.6 1 1469.3 63 16 R.KAETIQADTPALSLIAETVEDM@VK.KHsNut2Med_Ti_04.010002.010002.2 2.4827 0.2733 2716.31 2719.121 6346.1 1 229.9 29.2 1 R.KAETIQADTPALSLIAETVEDM@VKK.NHsNut2Med_Ti_03.014290.014290.3 6.9173 0.4918 2719.79 2719.121 5599.8 1 1053 37.5 14 R.KAETIQADTPALSLIAETVEDM@VKK.NHsNut2Med_Ti_02.018845.018845.3 4.6164 0.4233 2445.11 2446.772 5381.1 1 865.8 38.6 2 K.AETIQADTPALSLIAETVEDMVK.KHsNut2Med_Ti_02.018866.018866.2 5.0736 0.4916 2446.13 2446.772 8201.2 1 1173.4 52.3 4 K.AETIQADTPALSLIAETVEDMVK.KHsNut2Med_Ti_02.014810.014810.2 6.3583 0.5044 2460.03 2462.772 6407.3 1 1715.3 61.4 13 K.AETIQADTPALSLIAETVEDM@VK.KHsNut2Med_Ti_03.014942.014942.3 5.5821 0.3568 2465.18 2462.772 7703.8 1 1446.9 37.5 7 K.AETIQADTPALSLIAETVEDM@VK.KHsNut2Med_Ti_07.011781.011781.3 4.6577 0.4044 4652.93 4655.197 8741.4 1 577.3 18.8 8 K.NLPPASSPGYGM@TTGNNPM@SGTTTPTNTFPGGPITTLFNM@SM@SIK.DHsNut2Med_Ti_06.004430.004430.2 4.5264 0.4345 1599.73 1600.644 5561.2 1 1547.8 88.5 26 K.DRHESVGHGEDFSK.VHsNut2Med_Ti_06.003974.003974.3 4.2926 0.2929 1600.34 1600.644 5837.3 1 1676.6 57.7 12 K.DRHESVGHGEDFSK.VHsNut2Med_Ti_08.009251.009251.2 2.3465 0.1183 1243.21 1243.526 4572.8 4 437.2 77.8 1 K.NHPM@LM@NLLK.DHsNut2Med_Ti_01.010845.010845.2 4.7942 0.5343 1995.77 1998.113 8649.6 1 2153.8 70.6 16 K.DNPAQDFSTLYGSSPLER.QHsNut2Med_Ti_02.010391.010391.3 5.0854 0.3594 2000.54 1998.113 6015.8 1 1047.6 44.1 4 K.DNPAQDFSTLYGSSPLER.Q

* HsNut2Med_Ti_02.004077.004077.2 2.7812 0.1245 1027.81 1026.116 4948.8 1 943.4 87.5 2 R.MEICSGSNK.THsNut2Med_Ti_02.016436.016436.2 6.1797 0.4178 2449.39 2450.613 4802.5 1 1518.3 63 4 R.LSSSDSIGPDVTDILSDIAEEASK.L

* HsNut2Med_Ti_02.019238.019238.3 7.4177 0.4954 4130.36 4132.442 5640.2 1 2092.3 34 15 R.LSSSDSIGPDVTDILSDIAEEASKLPSTSDDCPAIGTPLR.D* HsNut2Med_Ti_03.007211.007211.2 5.0433 0.3941 1698.49 1700.852 7934.1 1 2062.5 76.7 7 K.LPSTSDDCPAIGTPLR.D* HsNut2Med_Ti_02.012423.012423.2 3.1523 0.2175 2280.79 2278.542 4004.7 1 544.2 50 1 K.GFASQALNTLGVPMLGGDNGETK.F* HsNut2Med_Ti_02.011097.011097.3 3.9347 0.2088 2292.71 2294.542 4900.5 1 1532.6 47.7 3 K.GFASQALNTLGVPM@LGGDNGETK.F* HsNut2Med_Ti_02.010864.010864.2 5.2152 0.3409 2296.43 2294.542 3771.3 1 533.5 52.3 8 K.GFASQALNTLGVPM@LGGDNGETK.F* HsNut2Med_Ti_07.010650.010650.2 4.3917 0.3196 2111.43 2112.347 6993.4 1 1296.5 63.2 8 K.FKGNNQADTVDFSIISVAGK.A* HsNut2Med_Ti_05.009986.009986.2 5.4263 0.3867 1838.35 1836.996 8137.1 1 1438.5 67.6 26 K.GNNQADTVDFSIISVAGK.A* HsNut2Med_Ti_06.009266.009266.3 3.1252 0.2051 2617.37 2617.937 5851.8 7 399.5 27 1 K.ALAPADLMEHHSGSQGPLLTTGDLGK.E* HsNut2Med_Ti_07.009882.009882.2 4.7601 0.3477 2634.97 2633.937 4877.1 1 672.5 46 4 K.ALAPADLM@EHHSGSQGPLLTTGDLGK.E* HsNut2Med_Ti_05.008842.008842.3 3.8966 0.305 2635.97 2633.937 6330.6 1 768.4 35 9 K.ALAPADLM@EHHSGSQGPLLTTGDLGK.E* HsNut2Med_Ti_07.010227.010227.3 4.3172 0.4217 2875.07 2875.226 6312.5 1 761.6 31.5 2 K.ALAPADLMEHHSGSQGPLLTTGDLGKEK.T* HsNut2Med_Ti_08.009495.009495.3 4.3949 0.4381 2890.28 2891.226 6261.4 2 472.3 28.7 3 K.ALAPADLM@EHHSGSQGPLLTTGDLGKEK.T* HsNut2Med_Ti_03.005763.005763.3 2.8493 0.1454 3003.26 3002.311 7052.4 2 516.5 24.1 1 K.EKTQKRVKEGNGTSNSTLSGPGLDSKPGK.R* HsNut2Med_Ti_03.005726.005726.2 4.1961 0.3575 2004.17 2004.118 6116.1 1 614.5 47.5 7 K.EGNGTSNSTLSGPGLDSKPGK.R* HsNut2Med_Ti_05.006220.006220.3 2.7919 0.1315 2160.02 2160.305 4155.8 1 468.3 33.3 1 K.EGNGTSNSTLSGPGLDSKPGKR.S

HsNut2Med_Ti_08.008514.008514.3 4.0742 0.4125 2117.54 2117.239 6611.8 1 566.3 31.2 4 K.SPSHSSSNRPFTPPTSTGGSK.SHsNut2Med_Ti_08.008526.008526.2 5.7797 0.5862 2118.51 2117.239 7102.6 1 550 47.5 2 K.SPSHSSSNRPFTPPTSTGGSK.SHsNut2Med_Ti_05.006950.006950.1 3.1657 0.3403 1389.44 1390.623 5488.1 1 475.5 53.8 4 R.SQTPPGVATPPIPK.IHsNut2Med_Ti_04.006812.006812.2 3.6449 0.2697 1390.17 1390.623 4757.6 1 637.2 69.2 3 R.SQTPPGVATPPIPK.IHsNut2Med_Ti_07.008606.008606.2 5.1784 0.47 2545.35 2546.726 3996.4 1 537.4 48 3 K.GTVM@VGKPSSHSQYTSSGSVSSSGSK.SHsNut2Med_Ti_08.008369.008369.3 5.0774 0.3068 2547.32 2546.726 7052.4 1 946.3 32 29 K.GTVM@VGKPSSHSQYTSSGSVSSSGSK.SHsNut2Med_Ti_06.004727.004727.3 3.4424 0.2052 1857.95 1857.885 6054 1 650.5 33.3 2 K.PSSHSQYTSSGSVSSSGSK.S

* HsNut2Med_Ti_07.006680.006680.2 4.9147 0.5357 1744.15 1745.717 6591.8 1 1983.6 79.4 6 K.SHHSHSSSSSSSASTSGK.M* HsNut2Med_Ti_07.006656.006656.3 4.3066 0.2101 1746.47 1745.717 5086 1 1332.2 51.5 3 K.SHHSHSSSSSSSASTSGK.M* HsNut2Med_Ti_02.004118.004118.2 4.5714 0.5159 1811.01 1811.873 8934.6 1 1885.3 67.6 4 K.LSSSM@YSSQGSSGSSQSK.N* HsNut2Med_Ti_06.005865.005865.3 3.9452 0.223 3324.35 3325.5 4601.7 1 397.8 25 2 K.LSSSM@YSSQGSSGSSQSKNSSQSGGKPGSSPITK.H* HsNut2Med_Ti_06.004626.004626.1 1.9696 0.1656 1530.68 1532.651 4055.5 1 195.7 50 1 K.NSSQSGGKPGSSPITK.H* HsNut2Med_Ti_06.004610.004610.2 5.074 0.36 1532.31 1532.651 5214.1 1 1160.5 76.7 9 K.NSSQSGGKPGSSPITK.H

HsNut2Med_Ti_06.003568.003568.1 2.7884 0.2593 1047.49 1048.098 4947.8 1 400.3 65 1 K.HGLSSGSSSTK.MHsNut2Med_Ti_08.008159.008159.2 3.2278 0.2492 1635.23 1633.761 5272.2 2 532.8 63.3 2 K.PNISPSHSRPPGGSDK.LHsNut2Med_Ti_08.008394.008394.2 2.8803 0.2328 1609.77 1610.91 4129.8 2 392.2 53.3 2 K.LASPM@KPVPGTPPSSK.A

* HsNut2Med_Ti_06.005720.005720.2 6.5531 0.5227 2013.97 2014.146 8157.2 1 2541.4 71.4 2 K.SPISSGSGGSHMSGTSSSSGMK.S* HsNut2Med_Ti_06.005692.005692.3 2.8669 0.1 2015.72 2014.146 6871.6 5 671.1 32.1 1 K.SPISSGSGGSHMSGTSSSSGMK.S* HsNut2Med_Ti_06.003746.003746.2 4.3197 0.4426 2044.23 2046.146 3256.3 1 331 52.4 1 K.SPISSGSGGSHM@SGTSSSSGM@K.S* HsNut2Med_Ti_07.002889.002889.3 3.197 0.2023 2046.26 2046.146 4548.1 1 1097.1 40.5 4 K.SPISSGSGGSHM@SGTSSSSGM@K.S* HsNut2Med_Ti_02.004760.004760.2 3.8118 0.3724 1282.23 1282.35 6685.1 1 1111.6 73.1 8 K.SSSGLGSSGSLSQK.T* HsNut2Med_Ti_02.004775.004775.1 3.1322 0.3878 1283.58 1282.35 4056.9 1 268.8 53.8 5 K.SSSGLGSSGSLSQK.T* HsNut2Med_Ti_05.005853.005853.3 6.1238 0.5717 3268.34 3269.271 4774.1 1 903 33.6 10 K.TPPSSNSCTASSSSFSSSGSSMSSSQNQHGSSK.G* HsNut2Med_Ti_05.005278.005278.3 6.0232 0.501 3284.63 3285.271 5897.9 1 677.8 26.6 9 K.TPPSSNSCTASSSSFSSSGSSM@SSSQNQHGSSK.G

HsNut2Med_Ti_07.009224.009224.2 3.3853 0.3534 1074.23 1072.29 3285.7 1 743.7 88.9 4 K.KPSLTAVIDK.L* HsNut2Med_Ti_06.007364.007364.3 6.8145 0.5378 3171.17 3173.368 5671.8 1 1084.5 33.1 11 K.HGVVTSGPGGEDPLDGQM@GVSTNSSSHPM@SSK.H* HsNut2Med_Ti_06.006573.006573.2 4.6085 0.4644 3172.25 3173.368 5267.7 1 446.4 35.5 2 K.HGVVTSGPGGEDPLDGQM@GVSTNSSSHPM@SSK.H* HsNut2Med_Ti_06.005270.005270.2 3.0914 0.2392 1190.75 1192.292 4817.1 2 545 75 4 K.HNMSGGEFQGK.R* HsNut2Med_Ti_05.003884.003884.2 3.2349 0.1795 1207.61 1208.292 4932.6 1 585.4 75 2 K.HNM@SGGEFQGK.R* HsNut2Med_Ti_02.003189.003189.2 2.8261 0.2609 1140.91 1141.177 3468.3 1 428.7 72.7 1 K.VSTSGSSVDSSK.K* HsNut2Med_Ti_07.009083.009083.2 4.7631 0.4473 2287.53 2288.581 4924.7 1 556.3 52.4 1 K.VTLQKPGESSGEGLRPQMASSK.N* HsNut2Med_Ti_07.008831.008831.2 4.2521 0.3248 2303.25 2304.581 3370.9 1 345.6 52.4 6 K.VTLQKPGESSGEGLRPQM@ASSK.N* HsNut2Med_Ti_04.005817.005817.2 4.649 0.4465 1718.17 1718.879 5874.9 1 1089.4 71.9 2 K.PGESSGEGLRPQMASSK.N* HsNut2Med_Ti_04.004979.004979.2 3.4908 0.3512 1734.15 1734.879 2894.6 1 204.8 53.1 2 K.PGESSGEGLRPQM@ASSK.N

HsNut2Med_Ti_03.002336.002336.2 2.695 0.3227 1320.95 1321.473 6025 4 475.8 58.3 2 K.NYGSPLISGSTPK.HHsNut2Med_Ti_03.006573.006573.1 2.8909 0.2049 1321.44 1321.473 3597.9 5 207.7 54.2 3 K.NYGSPLISGSTPK.H

* HsNut2Med_Ti_01.007811.007811.2 4.7722 0.4613 2298.07 2298.379 6579.7 1 876.7 54.8 2 K.SPAYTPQNLDSESESGSSIAEK.S* HsNut2Med_Ti_03.006771.006771.2 4.9713 0.4123 2469.83 2471.596 5739.1 1 900.1 52.4 2 K.SYQNSPSSDDGIRPLPEYSTEK.H* HsNut2Med_Ti_07.008829.008829.1 2.8239 0.2181 1285.34 1286.43 5626.2 1 441.4 60 2 K.SHSIKPESWSK.S* HsNut2Med_Ti_05.008072.008072.3 3.8305 0.2966 2570.48 2567.788 5872.9 8 442.1 28.3 1 K.SPISSDQSLSM@TSNTILSADRPSR.Lgi|28558975|re 42 354 64.10% 651 72890 7.4 U* HsNut2Med_Ti_02.005564.005564.2 2.5614 0.1208 1036.97 1037.121 5267.5 1 797.8 81.2 2 R.ISIESACEK.Q* HsNut2Med_Ti_06.009518.009518.2 4.1113 0.4506 2771.27 2773.091 6099.6 1 554.9 41.7 8 K.QVHEVGLDGTETYLPPLSM@SQNLAR.L* HsNut2Med_Ti_03.012393.012393.2 3.3078 0.2156 1519.27 1518.715 3997.1 1 960.4 86.4 4 K.FQPSLWPWDSVR.N* HsNut2Med_Ti_06.019239.019239.2 3.5389 0.3017 1970.65 1970.298 10606.2 1 1014.6 53.1 4 R.SALTEMCVLYDVLSIVR.D* HsNut2Med_Ti_05.016862.016862.2 5.5162 0.4011 1987.41 1986.298 6719.1 1 1436.3 68.8 67 R.SALTEM@CVLYDVLSIVR.D* HsNut2Med_Ti_07.013730.013730.3 4.8525 0.2352 1987.73 1986.298 8100.6 1 2364.4 51.6 8 R.SALTEM@CVLYDVLSIVR.D* HsNut2Med_Ti_05.008487.008487.2 3.8794 0.2995 1802.65 1804.112 6070.7 1 1016.9 66.7 10 K.KFM@TLDPVSQDALPPK.Q* HsNut2Med_Ti_05.008036.008036.3 4.0337 0.2274 1804.85 1804.112 4348.4 1 1191.9 55 2 K.KFM@TLDPVSQDALPPK.Q* HsNut2Med_Ti_08.009987.009987.2 3.1385 0.2687 3055.27 3055.56 5517.1 2 357 32.7 1 K.KFM@TLDPVSQDALPPKQNPQTLQLISK.K* HsNut2Med_Ti_02.008247.008247.2 3.2855 0.1431 1678.45 1675.938 5445.6 1 760.9 67.9 1 K.FM@TLDPVSQDALPPK.Q* HsNut2Med_Ti_05.009626.009626.3 4.5431 0.3337 2924.81 2927.386 5401.8 1 745.8 34 3 K.FM@TLDPVSQDALPPKQNPQTLQLISK.K* HsNut2Med_Ti_03.006747.006747.1 2.3374 0.0838 1269.53 1270.472 1967.9 1 132.4 70 1 K.QNPQTLQLISK.K* HsNut2Med_Ti_03.006746.006746.2 2.7946 0.2408 1272.11 1270.472 3575.5 6 408.5 80 1 K.QNPQTLQLISK.K* HsNut2Med_Ti_04.008498.008498.2 4.0017 0.2724 1085.11 1085.332 6789.8 1 1644.9 90 4 K.SLAGAAQILLK.G* HsNut2Med_Ti_04.008567.008567.1 1.8015 0.1212 1086.43 1085.332 9046.2 4 658 65 1 K.SLAGAAQILLK.G* HsNut2Med_Ti_03.004761.004761.2 3.107 0.1539 1446.19 1446.56 6469.3 1 865.9 72.7 1 K.SVTENQENKLQR.D* HsNut2Med_Ti_07.019338.019338.2 2.8222 0.175 1725.87 1727.96 6434.4 2 305.8 46.7 1 R.SAGSLFPHHGTFEVIK.N* HsNut2Med_Ti_03.011576.011576.3 4.9065 0.2047 2750.63 2752.057 5590.7 1 969.9 35.9 7 K.KIPEDYCPLDVQIPSDLEGSAYIK.V* HsNut2Med_Ti_03.011222.011222.2 4.9472 0.3378 2754.65 2752.057 7045.9 1 717.6 43.5 5 K.KIPEDYCPLDVQIPSDLEGSAYIK.V* HsNut2Med_Ti_02.011997.011997.2 5.7663 0.3309 2624.27 2623.882 5883.8 1 764.7 50 8 K.IPEDYCPLDVQIPSDLEGSAYIK.V* HsNut2Med_Ti_02.012021.012021.3 3.9247 0.1234 2625.53 2623.882 6839.6 1 1079.7 35.2 1 K.IPEDYCPLDVQIPSDLEGSAYIK.V* HsNut2Med_Ti_02.011174.011174.2 3.3095 0.1966 1590.31 1588.801 6367.9 1 743 60.7 2 K.QAPDIGDLGTVNLFK.R* HsNut2Med_Ti_08.008255.008255.2 2.7558 0.2243 1252.51 1253.403 5135.9 1 687.2 85 1 K.SKPGSPHWQTK.L* HsNut2Med_Ti_02.007116.007116.2 2.6272 0.2091 1258.13 1259.455 7847.2 1 854.7 75 1 K.LEAAQNVLLCK.E* HsNut2Med_Ti_02.007088.007088.2 2.9291 0.0996 964.01 964.109 6393.5 1 1115.9 92.9 1 K.EIFAQLSR.E* HsNut2Med_Ti_06.011638.011638.2 4.3707 0.3592 2715.27 2715.001 7377 1 639.4 45.7 8 K.NQIISQPFPSLQLSISLCHSSNDK.K* HsNut2Med_Ti_07.011194.011194.2 3.5828 0.2791 2842.31 2843.175 6740.1 4 227.6 33.3 2 K.NQIISQPFPSLQLSISLCHSSNDKK.S* HsNut2Med_Ti_07.011666.011666.3 3.6422 0.2676 2400.98 2400.709 6411.4 1 1195.5 45.8 3 K.QCPEDHLYVLEHNLHLLIR.E* HsNut2Med_Ti_07.011992.011992.2 2.9456 0.0802 2402.31 2400.709 3586 1 162.4 41.7 1 K.QCPEDHLYVLEHNLHLLIR.E* HsNut2Med_Ti_03.009543.009543.3 4.2007 0.2974 2608.7 2605.861 2960.4 1 380 34.8 1 R.LSGPQAFDKNEINSLQSSEGLLEK.I* HsNut2Med_Ti_03.007918.007918.2 4.9323 0.3233 1663.51 1661.806 9806 1 3134.5 89.3 7 K.NEINSLQSSEGLLEK.I* HsNut2Med_Ti_02.006120.006120.1 2.7846 0.2809 1077.45 1076.195 3664.7 1 257.9 60 2 R.AAATIDSLASR.I* HsNut2Med_Ti_03.009370.009370.3 6.0404 0.3933 2572.73 2573.778 5172 1 1360.4 42.9 3 R.IEDPQIQAHWSNINDVYESSVK.V* HsNut2Med_Ti_04.008268.008268.2 5.5229 0.4111 2573.95 2573.778 4369.2 1 696.3 57.1 5 R.IEDPQIQAHWSNINDVYESSVK.V* HsNut2Med_Ti_03.007001.007001.1 2.6431 0.2136 1538.42 1539.735 5679.9 1 582.8 62.5 3 K.VLITSQGYEQICK.S* HsNut2Med_Ti_02.006968.006968.2 4.2997 0.4033 1540.27 1539.735 8616.8 1 1928.3 83.3 21 K.VLITSQGYEQICK.S* HsNut2Med_Ti_05.011066.011066.2 5.0766 0.2098 1612.01 1611.882 7845.7 1 1893.5 80.8 88 K.SIQLQLNIGVEQIR.V* HsNut2Med_Ti_08.011461.011461.3 5.792 0.3827 3757.01 3758.241 8942.8 1 1114 26.6 14 R.VITLSYQEQELQDFLLSQM@SQHQVHAVQQLAK.V* HsNut2Med_Ti_08.012269.012269.3 7.4199 0.5673 4203.62 4205.733 9083.8 1 2297.4 31.2 38 K.VM@GWQVLSFSNHVGLGPIESIGNASAITVASPSGDYAISVR.N* HsNut2Med_Ti_05.012986.012986.2 4.8819 0.2164 2163.35 2162.575 5563.4 1 828.1 58.8 7 R.NFVYKM@ELLM@SALSPCLL.-* HsNut2Med_Ti_01.011722.011722.2 3.273 0.1896 1509.97 1510.812 6085 1 1303.6 75 3 K.M@ELLM@SALSPCLL.-* HsNut2Med_Ti_01.011831.011831.1 2.7259 0.116 1510.28 1510.812 6928.8 1 594.3 58.3 3 K.M@ELLM@SALSPCLL.-gi|41281496|re 72 587 63.10% 989 110305 6.9 U MEDIATOR_TRAP100 thyroid hormone receptor-associated protein; thyroid hormone receptor-associated protein (100 kDa) [Homo sapiens]

HsNut2Med_Ti_07.018816.018816.3 6.3134 0.4404 3457.07 3456.035 10090.8 1 1977.1 32.3 10 K.GATWDILNLADALLEQAM@IGPSPNPLILSYLK.YHsNut2Med_Ti_04.009099.009099.2 5.4346 0.5329 2051.19 2052.348 8231.5 1 2538.5 75 9 K.YAISSQM@VSYSSVLTAISK.FHsNut2Med_Ti_03.010854.010854.3 4.6434 0.3743 2053.94 2052.348 7996.4 1 1880.8 47.2 2 K.YAISSQM@VSYSSVLTAISK.FHsNut2Med_Ti_05.000188.000188.3 3.9175 0.3339 2819.78 2820.144 5954.4 1 473.2 27.1 2 K.YAISSQM@VSYSSVLTAISKFDDFSR.DHsNut2Med_Ti_03.015483.015483.2 5.4762 0.4415 2149.75 2148.384 8139.1 1 1813.6 71.9 52 R.DLCVQALLDIM@DM@FCDR.LHsNut2Med_Ti_02.005913.005913.1 2.1712 0.1387 1107.44 1108.169 4289.3 6 343.7 68.8 1 K.AEECIGLCR.AHsNut2Med_Ti_08.012263.012263.2 3.6604 0.2698 1293.55 1293.598 7312.5 1 1510 85 2 R.ALLSALHWLLR.CHsNut2Med_Ti_04.005854.005854.1 2.7796 0.3331 1498.45 1499.664 3387.9 3 253.8 57.1 1 R.LREGLEAGTPAAGEK.QHsNut2Med_Ti_04.005840.005840.2 5.0421 0.4351 1502.11 1499.664 7231.8 1 2164.1 85.7 8 R.LREGLEAGTPAAGEK.QHsNut2Med_Ti_02.004761.004761.1 2.4315 0.1764 1229.36 1230.317 2514.1 2 161.3 54.2 2 R.EGLEAGTPAAGEK.QHsNut2Med_Ti_02.004862.004862.2 3.3983 0.1613 1233.01 1230.317 6144.8 3 861.1 75 2 R.EGLEAGTPAAGEK.QHsNut2Med_Ti_03.010965.010965.3 2.9869 0.2108 1815.05 1815.033 2841.5 1 498.9 41.7 1 K.LEEASSWTAIEHSLLK.LHsNut2Med_Ti_05.011324.011324.2 4.4233 0.3073 1817.31 1815.033 8637.4 1 1413.2 66.7 8 K.LEEASSWTAIEHSLLK.LHsNut2Med_Ti_03.009700.009700.1 2.945 0.3003 1537.9 1538.787 3930.6 1 311.1 61.5 2 K.LGEILANLSNPQLR.SHsNut2Med_Ti_03.009760.009760.3 3.2277 0.1294 1538.63 1538.787 5436.5 5 928.9 51.9 1 K.LGEILANLSNPQLR.SHsNut2Med_Ti_03.009746.009746.2 4.5089 0.2774 1540.67 1538.787 5911.4 1 1432.9 80.8 4 K.LGEILANLSNPQLR.SHsNut2Med_Ti_03.002397.002397.2 2.8482 0.1519 1263.23 1263.359 5849.4 2 491.9 65 2 R.SQAEQCGTLIR.SHsNut2Med_Ti_07.009618.009618.2 3.7101 0.2765 1742.39 1742.019 3333.9 4 233.3 60.7 1 R.SIPTM@LSVHAEQM@HK.THsNut2Med_Ti_06.018128.018128.3 7.4107 0.4959 3592.22 3592.159 8268 1 1486.8 29.7 67 K.TGFPTVHAVILLEGTM@NLTGETQSLVEQLTM@VK.RHsNut2Med_Ti_08.012516.012516.2 2.8333 0.1155 1868.43 1869.276 3650.8 1 364.3 57.1 1 R.M@QHIPTPLFVLEIWK.AHsNut2Med_Ti_02.009794.009794.2 5.4901 0.3048 1879.17 1880.068 6915.5 1 2632.2 84.4 2 K.ACFVGLIESPEGTEELK.WHsNut2Med_Ti_03.010976.010976.2 2.7202 0.1279 1016.21 1014.212 3548.6 1 430.2 92.9 2 K.WTAFTFLK.IHsNut2Med_Ti_02.017330.017330.2 5.4395 0.4167 1848.23 1849.013 7064.6 1 2086 82.1 37 K.DFTEDVNCAFEFLLK.LHsNut2Med_Ti_02.017296.017296.3 4.7833 0.2845 1849.13 1849.013 9470.7 1 1611.2 50 3 K.DFTEDVNCAFEFLLK.LHsNut2Med_Ti_03.007650.007650.1 1.6556 0.2758 1590.58 1591.821 1901.9 4 59 50 1 K.QGLLSEASVNNLM@AK.RHsNut2Med_Ti_03.007634.007634.2 4.688 0.4479 1590.67 1591.821 5861.6 1 1622.4 82.1 2 K.QGLLSEASVNNLM@AK.RHsNut2Med_Ti_02.009888.009888.2 4.8065 0.3472 1778.55 1781.022 5119.5 1 877 66.7 9 K.SGENANIQPNIQLILR.AHsNut2Med_Ti_07.013512.013512.3 4.6706 0.3514 1595.3 1595.898 5832 1 2148.1 58.3 2 K.SPEGLLGVLGHMLSGK.SHsNut2Med_Ti_07.013509.013509.2 4.9251 0.4183 1596.29 1595.898 8146.3 1 2240.2 80 2 K.SPEGLLGVLGHMLSGK.SHsNut2Med_Ti_08.011339.011339.3 4.2441 0.3029 1611.86 1611.898 4429.2 1 990.5 50 2 K.SPEGLLGVLGHM@LSGK.SHsNut2Med_Ti_08.011322.011322.2 4.6323 0.3106 1612.07 1611.898 5246.4 1 1011.9 73.3 2 K.SPEGLLGVLGHM@LSGK.SHsNut2Med_Ti_04.009628.009628.1 2.9587 0.2197 1314.46 1315.553 7541.1 5 489.5 53.8 2 K.SLDLLLAAAAATGK.LHsNut2Med_Ti_03.001916.001916.2 4.1137 0.2823 1316.05 1315.553 4073 1 680.9 73.1 23 K.SLDLLLAAAAATGK.LHsNut2Med_Ti_05.009491.009491.2 5.2472 0.449 2110.13 2109.297 8683.1 1 1920.1 73.5 4 R.KFINLNEFTTYGSEESTK.PHsNut2Med_Ti_08.010199.010199.3 4.6365 0.2882 2618.54 2619.891 8806.9 1 936.1 33 2 R.KFINLNEFTTYGSEESTKPASVR.AHsNut2Med_Ti_02.010268.010268.2 6.197 0.4462 1983.13 1981.123 9219.1 1 3090.4 87.5 4 K.FINLNEFTTYGSEESTK.PHsNut2Med_Ti_04.008553.008553.2 5.0368 0.3851 2490.29 2491.717 7569.1 1 1026.4 52.4 9 K.FINLNEFTTYGSEESTKPASVR.AHsNut2Med_Ti_05.010028.010028.3 4.2709 0.2326 2492.3 2491.717 4931.5 1 788.2 39.3 2 K.FINLNEFTTYGSEESTKPASVR.A

MEDIATOR_TRAP80 {cofactor required for Sp1 transcriptional activation, subunit 6, 77kDa; thyroid hormone receptor-associated protein, 80-KD subunit; vitamin D receptor interacting protein 80-kD}

HsNut2Med_Ti_02.011192.011192.3 6.5104 0.4233 2510.48 2508.744 4687.1 1 1498.3 48.8 3 R.TGAEVPFFETWM@QTCM@PEEGK.IHsNut2Med_Ti_02.011084.011084.2 5.325 0.425 2510.93 2508.744 6799.3 1 918.4 52.5 9 R.TGAEVPFFETWM@QTCM@PEEGK.IHsNut2Med_Ti_07.009165.009165.2 2.6994 0.2518 1797.15 1797.974 7851.2 1 570.4 53.6 1 K.ILNPDHPCFRPDSTK.VHsNut2Med_Ti_08.011165.011165.3 5.9004 0.4449 3427.07 3429.837 5325.8 1 904 32.8 2 K.ILNPDHPCFRPDSTKVESLVALLNNSSEM@K.LHsNut2Med_Ti_02.009959.009959.2 3.6454 0.2113 1633.71 1634.885 6137.8 1 846 64.3 2 K.VESLVALLNNSSEMK.LHsNut2Med_Ti_02.009866.009866.3 4.2968 0.286 1650.32 1650.885 6129.3 1 2804.9 64.3 2 K.VESLVALLNNSSEM@K.LHsNut2Med_Ti_06.010451.010451.2 3.5711 0.2897 1651.73 1650.885 7256.1 1 1086.7 67.9 6 K.VESLVALLNNSSEM@K.LHsNut2Med_Ti_08.013602.013602.3 3.6623 0.4017 1911.47 1912.218 5619.3 3 619.3 39.1 2 K.VCSLAVCAVAWLVAHVR.MHsNut2Med_Ti_08.013607.013607.2 4.7634 0.4858 1913.31 1912.218 8336.7 1 1765.9 68.8 1 K.VCSLAVCAVAWLVAHVR.MHsNut2Med_Ti_03.011360.011360.2 5.1809 0.3688 2128.23 2128.349 8026.7 1 1819.6 67.6 8 R.QLAGPLFSENTLQFYNER.VHsNut2Med_Ti_03.011444.011444.3 5.3289 0.2783 2128.73 2128.349 7192.1 1 2200.5 51.5 2 R.QLAGPLFSENTLQFYNER.VHsNut2Med_Ti_02.006932.006932.2 5.4812 0.3524 1626.25 1623.829 9958.4 1 3346.3 92.3 7 R.M@CADVLQQTATQIK.FHsNut2Med_Ti_04.009293.009293.2 4.4856 0.3842 2080.97 2080.406 6639.1 1 1015.3 64.7 12 K.FPSTGVDTM@PYWNLLPPK.RHsNut2Med_Ti_02.009056.009056.2 2.3642 0.1822 1035.67 1036.213 6417.9 1 698.4 81.2 2 K.EVLTDIFAK.VHsNut2Med_Ti_07.009441.009441.2 4.2986 0.3538 1539.61 1540.778 4034 1 574.2 65.4 2 K.WHSLM@DPPGTALAK.LHsNut2Med_Ti_07.009508.009508.1 2.6813 0.3314 1540.47 1540.778 3758.2 2 227.1 50 4 K.WHSLM@DPPGTALAK.LHsNut2Med_Ti_08.010441.010441.2 4.1105 0.2906 1611.37 1609.788 5966.6 1 1511.4 80.8 2 K.LAVWCALSSYSSHK.GHsNut2Med_Ti_08.014035.014035.3 3.8686 0.2054 2675.09 2673.942 9557.2 1 1003.9 33.3 5 K.RHREDIEDYISLFPLDDVQPSK.LHsNut2Med_Ti_07.012116.012116.2 5.4688 0.4224 2517.41 2517.755 8808.2 1 1685.6 60 83 R.HREDIEDYISLFPLDDVQPSK.LHsNut2Med_Ti_05.016244.016244.3 5.6302 0.2769 2517.56 2517.755 4847.6 1 1261 45 49 R.HREDIEDYISLFPLDDVQPSK.LHsNut2Med_Ti_07.012753.012753.3 4.7122 0.2744 2933.63 2934.294 6333.6 1 661.5 33.7 1 R.HREDIEDYISLFPLDDVQPSKLM@R.LHsNut2Med_Ti_02.014487.014487.2 6.0366 0.4354 2226.17 2224.426 5985.1 1 1008.6 61.1 2 R.EDIEDYISLFPLDDVQPSK.LHsNut2Med_Ti_02.007365.007365.2 5.3725 0.4408 1948.09 1948.051 8681.8 1 1712.2 67.6 4 R.LLSSNEDDANILSSPTDR.SHsNut2Med_Ti_06.006338.006338.2 3.7076 0.3804 1870.07 1869.076 3382.9 1 362.5 56.2 7 R.SM@SSSLSASQLHTVNM@R.DHsNut2Med_Ti_08.015668.015668.3 4.0498 0.3069 1716.5 1717.105 4914.7 3 1083.6 48.3 3 R.VLANLFLLISSILGSR.THsNut2Med_Ti_08.015965.015965.2 4.49 0.3563 1716.53 1717.105 7136.4 1 1878.4 80 18 R.VLANLFLLISSILGSR.THsNut2Med_Ti_08.012183.012183.3 3.1507 0.1069 2983.52 2984.201 7822.7 5 476.4 25 1 R.TAGPHTQFVQWFMEECVDCLEQGGR.GHsNut2Med_Ti_06.011495.011495.3 6.5785 0.4443 3001.97 3000.201 9803.9 1 2385.8 38.5 3 R.TAGPHTQFVQWFM@EECVDCLEQGGR.GHsNut2Med_Ti_03.015131.015131.2 5.3587 0.3332 1885.75 1884.241 5023.6 1 1539.4 78.1 2 R.GSVLQFMPFTTVSELVK.VHsNut2Med_Ti_02.013574.013574.2 5.5857 0.393 1900.37 1900.241 6248.8 1 2326.8 81.2 26 R.GSVLQFM@PFTTVSELVK.VHsNut2Med_Ti_03.013612.013612.3 3.3386 0.106 1900.64 1900.241 7311.1 2 922.5 39.1 1 R.GSVLQFM@PFTTVSELVK.VHsNut2Med_Ti_03.012795.012795.3 3.5846 0.0937 1467.98 1467.792 4759.3 9 1098 57.7 1 K.VVLAITDLSLPLGR.QHsNut2Med_Ti_04.009956.009956.2 4.6721 0.3978 1468.25 1467.792 4458.3 1 1196.2 84.6 18 K.VVLAITDLSLPLGR.QHsNut2Med_Ti_03.012855.012855.1 3.3771 0.1881 1468.8 1467.792 4040.1 1 418.1 61.5 8 K.VVLAITDLSLPLGR.Q

gi|8923053|ref| 14 142 63.00% 208 23651 6.5 U MEDIATOR_p28b {hypothetical protein FLJ20045} [Homo sapiens] * HsNut2Med_Ti_06.011655.011655.3 5.4238 0.3268 2842.61 2841.234 4270.7 1 606.3 38.6 4 R.LRGLCDNM@EPETFLDHEM@VFLLK.G* HsNut2Med_Ti_06.012260.012260.3 3.3773 0.2877 2569.28 2571.887 4704.7 2 456.9 33.8 2 R.GLCDNM@EPETFLDHEM@VFLLK.G* HsNut2Med_Ti_05.012069.012069.2 3.9881 0.3015 2571.97 2571.887 4371.1 1 196.2 37.5 6 R.GLCDNM@EPETFLDHEM@VFLLK.G* HsNut2Med_Ti_03.006986.006986.2 3.1717 0.1611 1103.43 1103.266 6260.8 1 1094.2 83.3 1 K.GQQASPFVLR.A* HsNut2Med_Ti_08.009188.009188.2 2.1161 0.1387 907.59 908.051 3528.1 8 484.6 92.9 1 R.AGAPWHLR.Y* HsNut2Med_Ti_02.004788.004788.1 1.7105 0.1489 1153.23 1154.281 2764.7 4 171.1 61.1 1 R.YLGQPEM@GDK.N* HsNut2Med_Ti_02.004773.004773.2 3.0075 0.1174 1154.99 1154.281 4200.4 3 773.5 83.3 2 R.YLGQPEM@GDK.N* HsNut2Med_Ti_02.014058.014058.3 6.28 0.3788 2496.5 2496.734 11691.6 1 2697.9 45 8 R.NCVDIATSENLTDFLM@EM@GFR.M* HsNut2Med_Ti_02.013371.013371.2 6.2647 0.4693 2498.43 2496.734 9722.1 1 2293.1 65 76 R.NCVDIATSENLTDFLM@EM@GFR.M* HsNut2Med_Ti_07.011565.011565.3 5.602 0.3561 3473.09 3470.833 10047.6 1 1379.5 30.4 10 R.NCVDIATSENLTDFLM@EM@GFRM@DHEFVAK.G* HsNut2Med_Ti_03.004476.004476.1 1.8558 0.1869 992.23 993.123 2230 1 173.1 71.4 2 R.M@DHEFVAK.G* HsNut2Med_Ti_03.005036.005036.2 2.8609 0.1544 992.55 993.123 4758.8 1 803.4 92.9 4 R.M@DHEFVAK.G* HsNut2Med_Ti_05.016733.016733.3 5.3498 0.4214 3900.38 3899.373 5376.5 1 536.4 23.6 23 R.ILVPGNTDSTEALSLSYLVELSVVAPAGQDM@VSDDM@K.N* HsNut2Med_Ti_08.010248.010248.2 4.1636 0.3438 1664.61 1666.961 9149.8 1 1157.8 69.2 2 K.NFAEQLKPLVHLEK.Igi|31652218|re 12 110 62.90% 140 16480 5 U MEDIATOR_Surf5a {surfeit 5 isoform a; surfeit locus protein 5} gi|5803183|ref|NP_006743.1| [Homo sapiens] gi|5803183|ref| 12 110 62.90% 140 16480 5 U MEDIATOR_Surf5a {surfeit 5 isoform a; surfeit locus protein 5} [Homo sapiens]

HsNut2Med_Ti_02.006368.006368.1 2.8833 0.1428 1096.36 1096.225 3347.5 1 252.3 75 2 K.ETLLQSYNK.RHsNut2Med_Ti_02.011376.011376.2 2.8748 0.0854 1311.09 1311.536 6714.1 5 617.4 70 1 K.SIMDNFTEIIK.THsNut2Med_Ti_02.008728.008728.2 3.6499 0.2666 1325.81 1327.536 5468.9 2 876.8 85 3 K.SIM@DNFTEIIK.THsNut2Med_Ti_03.004902.004902.1 2.7726 0.2372 1377.41 1377.494 5335 1 320.5 54.5 2 K.TAKIEDETQVSR.AHsNut2Med_Ti_03.004703.004703.2 3.6087 0.1999 1380.19 1377.494 7660.8 1 1461.5 81.8 2 K.TAKIEDETQVSR.AHsNut2Med_Ti_03.004100.004100.2 4.1375 0.3922 1694.19 1694.774 5681.4 1 765.6 69.2 4 R.ATQGEQDNYEM@HVR.AHsNut2Med_Ti_03.013382.013382.2 5.8884 0.3716 2120.25 2120.37 6368.8 1 1700.7 73.5 38 K.QFLILNDFPSVNEAIDQR.NHsNut2Med_Ti_03.013283.013283.3 5.8837 0.2796 2121.53 2120.37 9232.4 1 3866 57.4 6 K.QFLILNDFPSVNEAIDQR.NHsNut2Med_Ti_08.013321.013321.3 3.4441 0.1834 2981.9 2979.31 10337.3 1 878.3 28.3 1 R.KLITLRDEISIDLYELEEEYYSSR.YHsNut2Med_Ti_07.013784.013784.3 4.7407 0.3322 2852.18 2851.136 6650 1 780.5 31.8 8 K.LITLRDEISIDLYELEEEYYSSR.YHsNut2Med_Ti_08.013393.013393.2 4.7421 0.3649 2853.59 2851.136 5377.9 1 778.5 50 8 K.LITLRDEISIDLYELEEEYYSSR.YHsNut2Med_Ti_06.014673.014673.2 6.0111 0.4049 2256.65 2254.366 12505.5 1 2690.1 67.6 35 R.DEISIDLYELEEEYYSSR.Y

gi|13097162|gb 14 129 60.00% 135 15688 6.2 U MEDIATOR_Nut2 {MGC5309 protein} [Homo sapiens] * HsNut2Med_Ti_04.006303.006303.1 3.2587 0.2325 1296.38 1297.41 5747.8 1 914.3 77.8 5 K.FDHLEEHLEK.F* HsNut2Med_Ti_04.006296.006296.3 3.7378 0.1328 1297.37 1297.41 5771.4 1 1302 66.7 4 K.FDHLEEHLEK.F* HsNut2Med_Ti_04.008060.008060.2 3.1864 0.1349 1297.47 1297.41 4148.8 1 619.5 77.8 14 K.FDHLEEHLEK.F* HsNut2Med_Ti_04.008522.008522.2 5.6309 0.4771 2129.61 2131.393 5650.6 1 1353.1 71.1 23 R.QLGIIVSDFQPSSQAGLNQK.L* HsNut2Med_Ti_05.015995.015995.3 4.742 0.3561 3589.34 3589.083 8443.5 1 779.7 23.4 3 R.QLGIIVSDFQPSSQAGLNQKLNFIVTGLQDIDK.C* HsNut2Med_Ti_03.011564.011564.1 3.2076 0.2141 1475.53 1476.713 6883 1 871.7 66.7 4 K.LNFIVTGLQDIDK.C* HsNut2Med_Ti_03.011535.011535.2 5.288 0.2906 1478.79 1476.713 7486.1 1 2289.9 87.5 40 K.LNFIVTGLQDIDK.C* HsNut2Med_Ti_08.012113.012113.3 4.0152 0.204 2720.66 2718.007 8229.3 3 749.7 33.3 2 K.CRQQLHDITVPLEVFEYIDQGR.N* HsNut2Med_Ti_08.012296.012296.3 6.4669 0.4726 3562.28 3562.973 10446.1 1 1719.4 33 8 K.CRQQLHDITVPLEVFEYIDQGRNPQLYTK.E* HsNut2Med_Ti_05.000024.000024.2 4.2644 0.3764 2400.39 2401.681 5061.4 1 540.3 55.3 5 R.QQLHDITVPLEVFEYIDQGR.N* HsNut2Med_Ti_05.000009.000009.3 4.9487 0.3217 2401.49 2401.681 8299.4 1 1281.9 40.8 9 R.QQLHDITVPLEVFEYIDQGR.N* HsNut2Med_Ti_05.014726.014726.3 4.5949 0.3703 3243.71 3246.647 7637.8 3 517.9 25 10 R.QQLHDITVPLEVFEYIDQGRNPQLYTK.E* HsNut2Med_Ti_03.008253.008253.1 2.5929 0.1015 1030.46 1031.238 8636.3 8 719.7 75 1 K.SLLIQELSK.V* HsNut2Med_Ti_03.008240.008240.2 3.0225 0.0903 1030.73 1031.238 4917.1 1 1074.2 93.8 1 K.SLLIQELSK.Vgi|23320913|gb 11 44 59.90% 177 19951 10 U MEDIATOR_LCMR1_2 {lung cancer metastasis-related protein} [Homo sapiens]

HsNut2Med_Ti_03.011492.011492.3 2.6281 0.3424 2894.12 2895.152 2780.1 1 283.5 31 1 R.ELPGSTELTGSTNLITHYNLEQAYNK.FHsNut2Med_Ti_03.010354.010354.2 5.1878 0.4505 2895.19 2895.152 5342.7 1 331.4 36 14 R.ELPGSTELTGSTNLITHYNLEQAYNK.FHsNut2Med_Ti_04.010124.010124.3 3.0428 0.1199 2595.44 2596.918 3165.2 1 627.7 35.9 1 K.LSNFLPDLPGMIDLPGSHDNSSLR.SHsNut2Med_Ti_04.009452.009452.2 4.6588 0.3039 2611.49 2612.918 8113.7 1 976.4 47.8 11 K.LSNFLPDLPGM@IDLPGSHDNSSLR.SHsNut2Med_Ti_08.011381.011381.3 2.6993 0.1506 2612.09 2612.918 2756.8 2 291 28.3 1 K.LSNFLPDLPGM@IDLPGSHDNSSLR.SHsNut2Med_Ti_07.012652.012652.2 5.0823 0.4383 2677.73 2678.162 6866.6 1 1067.9 54.2 2 R.SLIEKPPILSSSFNPITGTMLAGFR.LHsNut2Med_Ti_07.011896.011896.3 3.5216 0.1063 2679.29 2678.162 10142.4 1 833.5 28.1 1 R.SLIEKPPILSSSFNPITGTMLAGFR.L

HsNut2Med_Ti_06.012942.012942.3 5.2964 0.4726 2691.89 2694.162 9809.7 1 1898.4 36.5 3 R.SLIEKPPILSSSFNPITGTM@LAGFR.LHsNut2Med_Ti_07.011835.011835.2 5.1296 0.4997 2692.61 2694.162 3781.6 1 214.6 39.6 4 R.SLIEKPPILSSSFNPITGTM@LAGFR.LHsNut2Med_Ti_07.008853.008853.1 2.7555 0.3228 1307.42 1308.446 5566.6 1 324 60 4 R.LHTGPLPEQCR.L

* HsNut2Med_Ti_07.008170.008170.2 3.7629 0.3667 2027.57 2029.106 4405.4 1 344.8 50 2 R.HSPDHPGM@GSSQASSSSSLR.-gi|4505941|ref| 59 197 58.80% 1174 133896 6.9 U* HsNut2Med_Ti_02.011163.011163.2 4.8828 0.4779 1986.53 1988.18 8640.1 1 1894.7 76.7 5 R.QQLDSFDEFIQM@SVQR.I* HsNut2Med_Ti_02.011151.011151.3 4.2855 0.2232 1987.91 1988.18 9608.6 4 1065.2 40 4 R.QQLDSFDEFIQM@SVQR.I* HsNut2Med_Ti_02.008638.008638.2 4.8806 0.4016 2801.75 2799.023 4985.4 1 793.1 52 3 R.IVEDAPPIDLQAEAQHASGEVEEPPR.Y* HsNut2Med_Ti_07.009873.009873.2 4.8909 0.3271 1837.65 1835.072 5763.9 1 1285.5 80.8 2 K.FEQIYLSKPTHWER.D* HsNut2Med_Ti_02.009549.009549.1 3.1754 0.4224 1597.47 1598.836 4069.9 1 513.7 65.4 4 R.NLTYSAPLYVDITK.T* HsNut2Med_Ti_03.003189.003189.2 3.7394 0.19 1455.29 1455.524 2941.6 1 305.7 72.7 1 K.EGEEQLQTQHQK.T* HsNut2Med_Ti_06.014536.014536.3 6.6404 0.4422 4035.32 4036.337 7025.7 1 945.4 27.2 16 R.STYCLLNGLTDRDLCELNECPLDPGGYFIINGSEK.V* HsNut2Med_Ti_02.012352.012352.2 4.7448 0.4297 2640.01 2641.823 5239.4 1 908 54.5 3 R.DLCELNECPLDPGGYFIINGSEK.V* HsNut2Med_Ti_03.006899.006899.1 2.2806 0.1114 1260.32 1261.479 2687 1 211.5 75 2 K.M@ATNTVYVFAK.K* HsNut2Med_Ti_03.006896.006896.2 3.8728 0.3232 1262.07 1261.479 3716.3 1 518 80 2 K.M@ATNTVYVFAK.K* HsNut2Med_Ti_06.011516.011516.2 2.2332 0.1587 1213.67 1214.494 2940.9 5 348.6 77.8 5 K.QEVPIIIVFR.A* HsNut2Med_Ti_05.007472.007472.2 3.4984 0.3787 1893.21 1894.116 5140.8 1 371.4 53.3 2 K.EM@LPHVGVSDFCETKK.A* HsNut2Med_Ti_08.009740.009740.2 3.4518 0.3136 1275.25 1273.496 6216.6 1 1098.4 88.9 2 K.AYFLGYM@VHR.L* HsNut2Med_Ti_04.008165.008165.2 2.6345 0.1044 827.97 827.058 3362.4 2 722 85.7 4 R.LLLAALGR.R* HsNut2Med_Ti_05.002134.002134.2 4.651 0.3387 1446.77 1446.776 6238.2 1 1768.6 87.5 9 R.LDLAGPLLAFLFR.G* HsNut2Med_Ti_03.010094.010094.2 2.9656 0.1068 1062.93 1063.239 5508.9 1 999.9 81.2 1 K.DFNLELAIK.T* HsNut2Med_Ti_02.007014.007014.1 2.4331 0.2998 1339.45 1340.435 5666.6 1 367.2 54.5 1 K.YSLATGNWGDQK.K* HsNut2Med_Ti_07.010259.010259.1 2.6657 0.2429 1245.52 1246.452 3528.3 1 234.7 70 1 R.LTFASTLSHLR.R* HsNut2Med_Ti_08.009999.009999.2 2.891 0.2311 1248.47 1246.452 6482.4 2 687.4 70 1 R.LTFASTLSHLR.R* HsNut2Med_Ti_08.008543.008543.2 2.8463 0.1779 912.91 913.068 3866.6 3 597.5 92.9 1 R.RLNSPIGR.D* HsNut2Med_Ti_08.010169.010169.3 5.7985 0.4635 2763.56 2762.122 6773.4 1 941 35.4 2 R.QLHNTLWGM@VCPAETPEGHAVGLVK.N* HsNut2Med_Ti_07.015800.015800.3 4.325 0.2891 4613.24 4613.206 7311.1 1 605.8 19.5 2 K.NLALM@AYISVGSQPSPILEFLEEWSM@ENLEEISPAAIADATK.I* HsNut2Med_Ti_07.010028.010028.2 3.6221 0.2931 1431.17 1430.659 7619.1 2 1127.1 77.3 2 K.IFVNGCWVGIHK.D* HsNut2Med_Ti_06.008750.008750.3 4.2953 0.2668 1709.87 1710.018 6223 1 2076.7 57.7 1 R.RQM@DIIVSEVSM@IR.D* HsNut2Med_Ti_02.009176.009176.2 4.5868 0.2827 1553.23 1553.83 5365.3 1 857.2 79.2 8 R.QM@DIIVSEVSM@IR.D* HsNut2Med_Ti_07.009760.009760.3 4.3886 0.1834 1241.72 1241.526 3976.4 9 1179.1 69.4 1 R.ICRPLLIVEK.Q* HsNut2Med_Ti_06.002090.002090.3 4.2021 0.3101 4570.49 4572.965 8367.4 1 739.7 21.1 1 R.EYNNYSWQDLVASGVVEYIDTLEEETVM@LAM@TPDDLQEK.E* HsNut2Med_Ti_08.010913.010913.3 5.2802 0.469 4204.94 4205.587 7027.8 1 561.2 22.1 2 K.EVAYCSTYTHCEIHPSM@ILGVCASIIPFPDHNQSPR.N* HsNut2Med_Ti_06.008924.008924.2 3.5462 0.2522 1535.25 1536.792 5392.6 1 861.6 70.8 2 K.QAMGVYITNFHVR.M* HsNut2Med_Ti_07.009814.009814.2 3.1142 0.3368 1553.27 1552.792 2539.7 8 170.2 62.5 3 K.QAM@GVYITNFHVR.M* HsNut2Med_Ti_04.007635.007635.3 2.7904 0.2056 1596.56 1595.854 6090.9 4 518.4 43.8 1 R.M@DTLAHVLYYPQK.P* HsNut2Med_Ti_07.010574.010574.3 4.8122 0.3902 2247.23 2247.66 7715.5 1 2273.8 50 2 R.MDTLAHVLYYPQKPLVTTR.S* HsNut2Med_Ti_07.010256.010256.2 4.7065 0.3686 2263.67 2263.66 4229.8 1 711.7 63.9 6 R.M@DTLAHVLYYPQKPLVTTR.S* HsNut2Med_Ti_06.009399.009399.3 3.5367 0.1378 2266.07 2263.66 6655.1 1 580.5 33.3 2 R.M@DTLAHVLYYPQKPLVTTR.S* HsNut2Med_Ti_07.014814.014814.3 4.2357 0.3431 3347.87 3346.74 7166.8 1 592.8 25.9 11 R.FRELPAGINSIVAIASYTGYNQEDSVIM@NR.S* HsNut2Med_Ti_03.014158.014158.3 4.4813 0.4208 3040.97 3043.376 10278.1 1 803.3 26.9 1 R.ELPAGINSIVAIASYTGYNQEDSVIM@NR.S* HsNut2Med_Ti_05.007239.007239.2 4.9453 0.4126 1709.59 1710.884 7504.3 1 1607.3 76.9 14 K.KGFDQEEVFEKPTR.E* HsNut2Med_Ti_05.009627.009627.3 4.3451 0.1735 1712.45 1710.884 5378.2 1 969.2 50 8 K.KGFDQEEVFEKPTR.E* HsNut2Med_Ti_03.006843.006843.2 4.0286 0.348 1584.17 1582.71 8565 1 1380.4 75 2 K.GFDQEEVFEKPTR.E* HsNut2Med_Ti_02.006028.006028.1 1.9766 0.2142 1001.8 1003.141 7658 3 835.4 72.2 1 R.VSGDDVIIGK.T* HsNut2Med_Ti_01.008108.008108.2 4.278 0.4608 1847.21 1847.93 4441.3 1 663.6 70 3 K.TVTLPENEDELESTNR.R* HsNut2Med_Ti_02.010798.010798.2 5.0635 0.5493 2229.27 2229.464 8985.9 1 2008.1 65.8 11 R.TSETGIVDQVM@VTLNQEGYK.F* HsNut2Med_Ti_05.010751.010751.3 3.9233 0.2619 2230.7 2229.464 8456.6 1 1016.8 38.2 1 R.TSETGIVDQVM@VTLNQEGYK.F* HsNut2Med_Ti_03.011850.011850.3 4.3307 0.3034 2855.9 2853.185 8060.5 1 875.6 31.2 2 R.QEDMPFTCEGITPDIIINPHAIPSR.M* HsNut2Med_Ti_07.010598.010598.1 3.5443 0.2975 1515.59 1516.763 2802.8 1 247.5 62.5 2 R.M@TIGHLIECLQGK.V* HsNut2Med_Ti_07.010571.010571.3 3.4132 0.1667 1516.49 1516.763 6942.5 2 802.8 47.9 1 R.M@TIGHLIECLQGK.V* HsNut2Med_Ti_02.007520.007520.2 4.7452 0.329 1777.15 1775.913 4774.3 1 964 71.9 4 K.GEIGDATPFNDAVNVQK.I* HsNut2Med_Ti_05.009189.009189.3 4.0975 0.1959 1551.47 1551.742 5874.6 2 1204.5 54.2 3 K.ISNLLSDYGYHLR.G* HsNut2Med_Ti_05.009196.009196.2 4.3875 0.2405 1554.47 1551.742 6237.4 1 1338.1 79.2 6 K.ISNLLSDYGYHLR.G* HsNut2Med_Ti_02.007241.007241.1 2.58 0.2978 1326.47 1327.439 2522 5 98.6 45.5 1 R.GNEVLYNGFTGR.K* HsNut2Med_Ti_03.008774.008774.2 4.0626 0.3439 1686.57 1687.936 5180.8 1 834.7 69.2 2 K.ITSQIFIGPTYYQR.L* HsNut2Med_Ti_08.009323.009323.2 3.1447 0.151 1137.55 1138.358 6172.5 1 655.6 72.2 1 R.ARGPIQILNR.Q* HsNut2Med_Ti_05.007973.007973.1 3.603 0.4282 1486.37 1487.624 4076.6 1 377.9 58.3 2 R.DCQIAHGAAQFLR.E* HsNut2Med_Ti_05.007895.007895.2 4.2126 0.3129 1486.77 1487.624 5748.9 1 1325.7 83.3 3 R.DCQIAHGAAQFLR.E* HsNut2Med_Ti_05.007934.007934.3 4.6462 0.3949 1489.64 1487.624 5034.9 1 1284.7 60.4 3 R.DCQIAHGAAQFLR.E* HsNut2Med_Ti_06.010731.010731.3 4.114 0.291 3180.95 3182.511 6145.7 1 416.8 26.9 1 R.ERLFEASDPYQVHVCNLCGIM@AIANTR.T* HsNut2Med_Ti_03.006116.006116.1 1.868 0.1146 817.48 816.976 3248.9 9 114.6 66.7 1 K.TQISLVR.M* HsNut2Med_Ti_03.010089.010089.2 4.3279 0.2682 1667.73 1669.008 6266 1 1513.5 80.8 7 K.LLFQELM@SM@SIAPR.M* HsNut2Med_Ti_04.008813.008813.3 3.8604 0.1861 1668.62 1669.008 8529.2 1 2637.7 59.6 2 K.LLFQELM@SM@SIAPR.Mgi|4505947|ref| 8 17 58.70% 172 19294 5.5 U PolII_Rpb7 {DNA directed RNA polymerase II polypeptide G; DNA directed RNA polymerase II 19 kda polypeptide} [Homo sapiens] * HsNut2Med_Ti_04.008492.008492.2 2.8879 0.2277 1266.41 1266.483 4701.9 3 539.3 75 1 R.YFGPNLLNTVK.Q* HsNut2Med_Ti_02.006420.006420.1 2.5566 0.3981 1341.61 1342.455 4600 1 330.7 59.1 3 K.LFTEVEGTCTGK.Y* HsNut2Med_Ti_02.006404.006404.2 4.6155 0.4012 1342.17 1342.455 3972.4 1 1009.2 90.9 2 K.LFTEVEGTCTGK.Y* HsNut2Med_Ti_07.012856.012856.2 3.6496 0.2465 2263.83 2262.614 8999.8 1 564.1 42.9 4 K.YGFVIAVTTIDNIGAGVIQPGR.G* HsNut2Med_Ti_08.010812.010812.3 6.2035 0.2697 2319.11 2317.737 10467 1 2951.3 45 2 K.AIVFRPFKGEVVDAVVTQVNK.V* HsNut2Med_Ti_02.006958.006958.2 3.1208 0.0847 1357.51 1358.535 4929.3 3 592 66.7 1 K.GEVVDAVVTQVNK.V* HsNut2Med_Ti_04.009356.009356.2 4.6094 0.225 1831.29 1831.104 4749.4 1 628 60 2 K.VGLFTEIGPM@SCFISR.H* HsNut2Med_Ti_04.006814.006814.2 4.2552 0.3533 2265.19 2266.41 6224.3 1 981.8 58.3 2 R.HSIPSEM@EFDPNSNPPCYK.Tgi|29428258|sp 9 58 58.50% 212 23222 6.9 U MEDIATOR_TRFP {TRFP_HUMAN TRF-proximal protein homolog} gi|30023851|ref|NP_004266.2| [Homo sapiens] gi|30023851|re 9 58 58.50% 212 23222 6.9 U MEDIATOR_TRFP {Trf-proximal; Trf (TATA binding protein-related factor)-proximal homolog (Drosophila)} [Homo sapiens]

HsNut2Med_Ti_02.008510.008510.1 2.5944 0.2651 1274.53 1274.46 4062.9 2 224.8 65 1 K.SVQQTVELLTR.KHsNut2Med_Ti_02.008558.008558.2 4.1233 0.0983 1276.53 1274.46 5854.9 1 1122.3 85 7 K.SVQQTVELLTR.KHsNut2Med_Ti_03.007311.007311.2 3.5828 0.3099 2806.29 2805.904 5206.9 2 199.1 32 5 K.QGTFCVDCETYHTAASTLGSQGQTGK.LHsNut2Med_Ti_08.013049.013049.3 7.2159 0.4243 4288.76 4289.854 9000.6 1 1579.6 28.6 17 K.LM@YVM@HNSEYPLSCFALFENGPCLIADTNFDVLM@VK.LHsNut2Med_Ti_02.008346.008346.2 2.8562 0.2579 1234.39 1236.376 3311.8 1 480.4 81.2 2 R.YQYCDFLVK.VHsNut2Med_Ti_04.004262.004262.2 3.6744 0.2718 1094.13 1092.256 3454.5 1 556.7 85 11 K.VGTVTM@GPSAR.GHsNut2Med_Ti_04.008142.008142.2 4.986 0.3931 2362.05 2362.635 4273.1 1 613.1 60.5 12 R.HDAVYGPADTM@VQYM@ELFNK.IHsNut2Med_Ti_04.008163.008163.3 6.2046 0.3317 2364.89 2362.635 7564.1 1 2288.4 50 2 R.HDAVYGPADTM@VQYM@ELFNK.IHsNut2Med_Ti_07.008844.008844.2 2.439 0.2232 1221.87 1224.449 6919.2 1 792.6 80 1 R.KQQQVPVAGIR.-

gi|5453930|ref| 7 23 58.40% 125 14523 5.1 U* HsNut2Med_Ti_02.006112.006112.2 3.1158 0.3041 1620.13 1620.75 5586.5 1 919.9 86.4 2 R.FCQECNNM@LYPK.E

PolII_Rpb9 {DNA directed RNA polymerase II polypeptide I; polymerase (RNA) II (DNA directed) polypeptide I (14.5kD); DNA directed RNA polymerase II 14.5 kda polypeptide} [Homo sapiens]

PolII_Rpb2 {DNA directe