7.01OverviewERIC T. KOOLUniversity of Rochester, NY, USA

6[90[0 DNA AS AN ORGANIC NATURAL PRODUCT 0

6[90[0[0 The Structure of DNA 06[90[0[1 The Functions of DNA 16[90[0[2 DNA is an Important Synthetic and Desi`n Problem 16[90[0[3 DNA Chemistry is Important to Medicine 3

6[90[1 IMPORTANT NATURAL PRODUCTS INTERACT WITH DNA 4

6[90[1[0 Natural Products that Bind Noncovalently 46[90[1[1 Natural Products that Dama`e DNA or Bind Covalently 46[90[1[2 Natural Products that Interact with DNA]Protein Interfaces 4

6[90[2 THE RELEVANCE OF MOLECULAR BIOLOGY TO CHEMISTRY 5

6[90[2[0 Molecular Biolo`y as a Branch of Chemistry 56[90[2[1 Molecular Biolo`y Offers Important Tools for Handlin` Lar`e Molecules 56[90[2[2 What Chemistry Offers to Molecular Biolo`y 5

6[90[3 WHAT IS COVERED IN THIS VOLUME 6

6[90[3[0 Physical Properties of DNA 66[90[3[1 Chemistry of DNA 66[90[3[2 Molecules that Bind to DNA 66[90[3[3 Techniques from Molecular Biolo`y 76[90[3[4 Topics not Covered 7

6[90[4 THE FUTURE OF DNA CHEMISTRY RESEARCH 7

6[90[4[0 Natural Products that Interact with DNA 86[90[4[1 DNA and Analo`ues as Dia`nostic Tools 86[90[4[2 DNA and Analo`ues as Therapeutic A`ents 86[90[4[3 DNA as a Molecular Scaffold 096[90[4[4 DNA as a Li`and 096[90[4[5 DNA as a Catalyst 006[90[4[6 DNA as a Computer 00

6[90[5 REFERENCES 01

6[90[0 DNA AS AN ORGANIC NATURAL PRODUCT

6[90[0[0 The Structure of DNA

The structure of DNA makes it abundantly clear that this molecule shares many things incommon with the smaller compounds that are normally regarded as natural products[ DNA containssubstructures*sugars and heterocycles*commonly found in many other classes of natural products"Figure 0#[ Of course\ it goes without saying that DNA is in fact a natural product\ that is\ an

0

1 Overview

organic compound that is biosynthesized in cells for a biological purpose[ Despite this\ however\chemists sometimes overlook DNA as a natural product[

Figure 0 The substructures of DNA\ a natural product found in all cells[

Why is this the case< It may be that large\ charged\ polymeric molecules tend to be overlooked indeference to less polar small organic compounds[ In part\ this is understandable\ considering thatcharged molecules require di}erent methods for synthesis and puri_cation than are commonlyused in most synthetically oriented laboratories "which commonly use solution!phase synthesis innonpolar solvents\ and puri_cation by silica column chromatography#[ In addition\ DNA is largeenough to make routine structural assignments by NMR spectroscopy a serious undertaking[Moreover\ it is usually made and handled on scales much smaller than the multimilligram scalescommon to most natural products chemists[

Nevertheless\ a very strong case can be made for DNA to be considered a natural product veryanalogous to smaller\ less polar molecules[ DNA is biosynthesized by a secondary metabolismprocess\ like most natural products[ Like such compounds\ DNA is biologically active\ and DNAinteracts with proteins to exert its biological activity[ Perhaps the one major di}erence betweenDNA and other commonly studied natural products is the fact that DNA is central to the existence\survival\ and replication of natural organisms\ while many other natural products are not[

6[90[0[1 The Functions of DNA

DNA is most widely thought of as the central storehouse of genetic information in each cell\ andof course\ that is a major function[ However\ DNA serves several other biological functions as well[It serves also as a replicable template for the copying of cells\ as a sca}old for the organization ofproteins\ and as a polyfunctional switch for control of gene expression[ In addition\ chemists haveused the DNA structure as a starting point for the design of many new molecules with functionsother than those listed for naturally occurring DNA[ Among these new functions are the ability toact as a ligand for nucleic acids and proteins and smaller molecules\ the ability to act as a sca}oldfor the organization of biological and nonnatural molecules\ and the ability to act as sensors andreporters of genetic sequences\ and even the ability to act as a computer[ Some of these functionsare discussed below or later in this volume[

6[90[0[2 DNA is an Important Synthetic and Design Problem

Because of the wide applications of DNA in chemistry\ biochemistry\ biology\ medicine\ andmaterials science\ there is broad motivation among chemists to develop and apply the chemistry ofDNA and related analogues[ Early in DNA research\ one of the _rst problems to be tackled wasthe task of assembling a string of nucleotides in a desired order and length[ Although biochemistsearly on isolated enzymes that could polymerize nucleotides "Figure 1#\0\1 it was di.cult or impossibleto manipulate the necessary templates in order to utilize these enzymes to make a desired sequenceof DNA[ Thus\ it was not until the development of synthetic methods for constructing speci_c DNAoligonucleotides on solid supports "Figure 2#2\3 that it became practical to study de_ned\ variedsequences with puri_ed molecules that were homogeneous in length[ Since the 0869s there has beenan explosion in the number of chemists working with DNA and in chemistries for manipulating it[

It can be argued that the ability to synthesize DNA oligonucleotides has been as important tothe molecular biology revolution as the development of bacterial cloning "see below#[ Also veryimportant to biochemistry has been the study of DNA damage and damaging agents\ and the studyof proteinÐDNA interactions^ these were also greatly aided by the ability to synthesize DNA[ Thus\

2Overview

Figure 1 The structure of the most well!studied and smallest DNA polymerase enzyme\ the Klenow fragmentof E[ coli DNA Polymerase I[0

Figure 2 The chemistry of the most commonly used synthetic method for assembly of DNA strands on asolid support[3

the progress made in developing the chemistry of natural DNA has been one of the most importantadvances in biologically!related _elds in the twentieth century[

One of the beautiful aspects of chemistry is that\ although chemists respect and admire naturalmolecules\ they are not con_ned to making and studying only those structures[ As a result\ the0879s and 0889s have seen the development of an extremely large number of synthetic modi_cations

3 Overview

to DNA|s natural structure "a few examples4Ð7 are shown in Figure 3#[ The wide variety of possibleapplications of DNA!like molecules has made this framework the subject of many molecular designprojects[ In addition\ the need for introducing many varied structural and functional groups intoDNA and DNA!like molecules has brought up the need for the development of new syntheticmethods as well[ It is also worth pointing out that DNA is not only synthesized on small scales forresearch purposes\ but oligodeoxynucleotides and analogues are also being tested in humans astherapeutic agents[8\09 This has led to the need for process!level synthetic chemistry aimed at theproduction of kilogram quantities of speci_c compounds[00\01 Overall\ then\ there are many strongmotivations for the continuing development of DNA chemistry[

Figure 3 Examples of published structural modi_cations made to the DNA backbone "natural structure isshown on the left#[4Ð7 All of the analogues shown can bind nucleic acids more tightly than natural DNA can[

6[90[0[3 DNA Chemistry is Important to Medicine

As mentioned above\ the development of chemistries for manipulating DNAs has been central tothe steady growth in biochemistry and molecular biology worldwide[ Scientists in these _elds havemade rapid progress in understanding biological processes such as cellular replication and cell death\gene expression\ cell signaling pathways\ and DNA damage and repair[ These processes "and defectsin them# have in many cases been linked directly to pathological conditions in humans[ For example\a speci_c chemical adduct of benzoðaŁpyrene to DNA "Figure 4# has been linked to a known\ speci_cpoint mutation in the p42 gene\02 which in turn has been suggested as the cause of lung cancer andother cancers[03\04 The ongoing rapid and large!scale sequencing of DNAs\05Ð08 also made possibleby the development of DNA chemistry\ has also led to the correlation of a growing set of geneticsequences to disease[ Many examples of inherited or acquired mutations are now directly linked tohuman disease states^ for example\ a number of mutated DNA sequences thought to be responsiblefor various leukemias are now known[19\10

Figure 4 The structure of an adduct of benzoðaŁpyrene diol epoxide to guanine in DNA[ Such adducts in thep42 gene have been directly linked to lung cancer[02

In the past one might have said that the chemistry of DNA is relevant only indirectly to medicine\in that it aids research in molecular biology or the understanding of biochemical processes\ but this

4Overview

is no longer the case[ There are currently a number of clinical trials underway to test the therapeuticpotential of synthetic DNA analogues in humans[8\09 In addition\ the US Food and Drug Admin!istration has begun to approve the use of medical diagnostic methods involving nonnaturallymodi_ed DNAs for the diagnosis of human disease[

6[90[1 IMPORTANT NATURAL PRODUCTS INTERACT WITH DNA

6[90[1[0 Natural Products that Bind Noncovalently

There are many natural products that have been shown to interact with DNA by formingnoncovalent complexes[ Although it may not always be true\ this binding may in many cases beresponsible for their biological activity[ The two major classes of known DNA!binding compoundsare the intercalators and the minor groove binders[ Note that intercalators are covered in thisvolume quite comprehensively in Chapter 6[01[ While there is no chapter speci_cally covering minorgroove binding\ a number of examples of molecules that bind in this fashion are discussed inChapters 6[02\ 6[03\ and 6[04[

6[90[1[1 Natural Products that Damage DNA or Bind Covalently

Many important natural products take advantage of their DNA binding properties to allow themto react with the DNA in some way[ For example\ it is common to _nd that electrophilic moleculesreact with nucleophilic sites in DNA bases\ or that free radicals generated from natural productswill react with deoxyribose moiety in DNA[ Some of these reaction mechanisms are amazing intheir elegance\ such as in the enediyene class of natural products "Chapter 6[04#[ Some such reactionmechanisms result in covalent attachment of a natural product to the DNA "at least temporarily#\while others never form a covalent bond with the DNA during their reaction with it[ DNA!damagingagents are comprehensively discussed here in Chapter 6[03\ and the chemical consequences of thisdamage are presented in Chapter 6[00[

6[90[1[2 Natural Products that Interact with DNA]Protein Interfaces

A relatively new concept in biologically in~uenced chemistry has been the idea of interfering atthe interface between two interacting proteins or between proteins and nucleic acids[ One of themost successful classes of natural products in this concept from a medical standpoint has been thetopoisomerase inhibitors[11\12 Topoisomerases are enzymes which alter the topology of DNA bybreaking and closing DNA strands\13Ð15 and topoisomerase inhibitors can exert biological activityby binding both a topoisomerase and its DNA target[ Compounds in this class such as adriamycin"also called doxorubicin# "Figure 5# are useful clinically in the treatment of cancer[16 Topoisomeraseinhibitors are discussed in detail in this volume "see Chapter 6[05#[

Figure 5 The structure of two clinically important topoisomerase inhibitors\ "a# adriamycin and "b# bisan!threne[ See Chapter 6[05 for details[

5 Overview

6[90[2 THE RELEVANCE OF MOLECULAR BIOLOGY TO CHEMISTRY

6[90[2[0 Molecular Biology as a Branch of Chemistry

The burgeoning _eld of molecular biology has arisen for two reasons] _rst\ there has been astrong desire to understand biological processes on a molecular basis[ This requires\ in part\identifying and purifying individual proteins and nucleic acids[ For example\ the observation thatcertain proteins are made at various times in a cell|s replication cycle led to the discovery of operatorsequences in DNAs and to the identi_cation of speci_c proteins such as repressors which canbind them and upregulate or downregulate gene expression[17\18 This led to the complete chemicalcharacterization of the sequence and structures of many such repressor molecules\ and even to therational design of altered DNA!binding proteins with new amino acid sequences[29\20 The _eld calledstructural biology\ which can be considered a subset of molecular biology\ strives further to identifythe three!dimensional structures of such large puri_ed molecules\ usually by X!ray crystallographicmethods or by solution!phase NMR spectroscopy[

To a chemist\ of course\ the ideas of puri_cation\ chemical and structural characterization\ andmolecular design are not new and in fact predate the _eld called molecular biology[ Since the goalsand motives are the same\ and since the molecules are in both cases organic\ it is reasonable toconsider molecular biology to be a branch of chemistry as well as a branch of biology[ It is alsotherefore rational to leave the descriptor {{chemistry|| in the name of the _eld^ hence the use of theterm {{chemical biology[||

6[90[2[1 Molecular Biology Offers Important Tools for Handling Large Molecules

Despite the fact that molecular biology is very closely related to chemistry\ it is worth noting thatscientists who call themselves molecular biologists have developed a variety of methods and toolsfor handling large molecules such as proteins and nucleic acids\ and some of these methods areparticularly useful for work with large charged compounds as opposed to small natural products[Chemists who are interested in working on such large molecules can bene_t greatly from using thetools of molecular biology[

For example\ there are a number of separation and puri_cation methods that are especially wellsuited to handling proteins and nucleic acids\ and which often do not have direct analogues in smallmolecule work^ among these methods are gel electrophoresis\ capillary electrophoresis\ isoelectricfocusing\ a.nity chromatography\ and size exclusion chromatography[ There are also a number ofsynthesis and preparation methods which are well worth learning if one is working with these largemolecules[ Among these methods are cloning and overexpression\ site!directed mutagenesis\ in vitrotranscription\ and ampli_cation via the polymerase chain reaction "PCR#[ There are importantidenti_cation methods as well[ Among these are direct protein and DNA sequencing\ Northern\Southern\ and Western blots\ and in situ hybridization[ Of course\ a number of analytical methodssuch as solution!phase synthesis\ solid!phase synthesis\ NMR spectroscopy\ HPLC\ and massspectrometry are useful for working with both small and large molecules[

Finally\ there are also a number of combinatorial methods developed for proteins and nucleicacids which are not readily applied to small molecules[ Among these methods are in vitro selectionand evolution of nucleic acids\ and phage display selection of peptides and proteins[

6[90[2[2 What Chemistry Offers to Molecular Biology

The above methods are examples of ways that chemistry can bene_t from molecular biology[This exchange of information is\ however\ a two!way street] there are also important ways in whichmolecular biologists can bene_t from the work of chemists[ For example\ chemists are probablymore likely than biologists to develop new analytical methods for molecular puri_cation\ identi!_cation\ and analysis\ and yet molecular biologists can and do apply these methods to their ownbiomolecules of interest[ Second\ chemists are much more likely to undertake the synthesis ofnonnatural molecules\ including close analogues of natural biomolecules[ Molecular biologists canuse such compounds as tools for their biological study\ for example\ as inhibitors or as probes ofmolecular interactions and mechanisms[

6Overview

6[90[3 WHAT IS COVERED IN THIS VOLUME

6[90[3[0 Physical Properties of DNA

Several chapters in this volume are focused on describing in detail the physical properties ofDNA[ Chapter 6[91 describes the thermodynamics and kinetics of DNA helix formation[ Thediscussion covers a number of important topics such as the forces which contribute to helixformation\ methods for measuring the thermodynamics of DNA helices\ and algorithms for pre!dicting the thermodynamics of DNA double helices a priori[ Also reviewed are the kinetics of DNAhelix formation and methods used to measure them[

Chapter 6[92 covers modern methods for the determination of DNA structure by nuclear magneticresonance spectroscopy[ Described are di}erent types of NMR experiments that are helpful inde_ning structure as well as practical descriptions on just what information can be gained fromthese experiments[

Because NMR methods are currently limited at times by the size of the molecules under inves!tigation\ we felt it prudent also to include a review of other methods for probing structure insolution[ Chapter 6[93 describes current chemical probing methods which are widely useful inexamination of DNA structure[ These methods are particularly useful in larger nucleic acids and inmapping protein!DNA interactions in solution[

6[90[3[1 Chemistry of DNA

Several chapters in this volume "Chapters 4Ð00# are concerned with the varied chemistry of DNA\ranging from its synthetic assembly to its destruction by chemically reactive species[ The _rst twoof these\ Chapters 6[94 and 6[95\ present in!depth coverage of modern methods for the chemicalsynthesis of DNA oligonucleotides as well as methods for attaching other moieties to such molecules\either during or after oligonucleotide construction[

Chapters 6Ð09 represent the most design!oriented chapters of the volume[ Chapter 6[96 describesnonnatural nucleoside analogues which have been widely useful in probing biochemical mechanismsand interactions[ Both the synthesis and the practical utility of these analogues are discussed[Chapters 6[97\ 6[98\ and 6[09 review the current state of the art in designing analogues of DNA forpractical applications[ Among the most important of these applications are the use of oligonucleotideanalogues as potential therapeutics in the downregulation of speci_c genes "e[g[\ antisense oligo!nucleotides#\ and the use of oligonucleotide analogues as molecular diagnostic tools for detectingand identifying disease!related genetic sequences[ Three di}erent chemical aspects of DNA arediscussed separately in these chapters[ Chapter 6[97 tackles the summary of a wide body of literatureon alterations made to the deoxyriboseÐphosphate diester backbone of DNA\ with a particular eyeto the synthesis and nucleic acid binding characteristics of each class of molecule[ Likewise\ Chapter6[98 covers the known alterations made to the nucleobases themselves\ again with emphasis on howthese changes a}ect binding properties[ Finally\ Chapter 6[09 covers structural alterations of ahigher order than backbone or bases[ Described here are topological alterations to the DNA chain"for example\ the e}ects of circular or knotted topologies# and the associated changes in properties\including ability to interact with proteins and nucleic acids[

The _nal chapter falling under the grouping of the chemistry of DNA is Chapter 6[00\ whichfocuses on the chemistry of DNA damage[ This is essentially a compendium of the reactions ofDNA in aqueous solution\ with a slant toward how this reactivity is relevant to disease states suchas cancer[ The chapter assesses various mechanisms by which reactive species such as electrophilesand radicals can initiate a cascade of further reactions in the DNA backbone or bases[ Peopleinterested in the mechanisms of DNA damage will also _nd Chapters 6[03 and 6[04 useful^ here aredescribed speci_c classes of natural products that bind and react with DNA[

6[90[3[2 Molecules that Bind to DNA

The second central grouping of chapters in this volume falls under the heading of molecules thatinteract with DNA[ Chapters 01 through 05 cover this topic in considerable detail[ Intercalators areamong the broadest classes of DNA!binding molecules^ these are reviewed in Chapter 6[01[ Anothervery broad class of DNA!binding compounds in nature are proteins and peptides^ although large

7 Overview

DNA!binding proteins are not explicitly discussed here "see below#\ smaller peptide motifs "includingsubdomains of active proteins# for binding DNA are covered in Chapter 6[02[

Another medically important class of natural products are those that bind DNA and do damageto the bases or backbone by their reactive nature[ Chapter 6[03 surveys the various known classesof reactive compounds and the reactions they promote on DNA substrate^ Chapter 6[04 focuses inmuch more detail on one fascinating class of DNA!reactive compounds\ the enediynes[

Finally\ also falling under the heading of DNA!binding molecules are those that interact with theinterface of proteins and DNA[ Chapter 6[05 covers topoisomerase inhibitors\ which are clinicallyimportant natural products[ Their mechanism of action and structureÐactivity relationships areexplored[

6[90[3[3 Techniques from Molecular Biology

The last two chapters of this volume are focused on techniques from molecular biology whichchemists are _nding increasingly useful[ Chapter 6[06 covers the rapidly expanding _eld of DNAswhich can act as ligands or as catalysts[ These have been discovered using in vitro selection methods\which typically involve making randomized libraries of molecules\ selecting for subsets which havedesirable properties\ and amplifying them prior to undergoing another round of selection[ Otherimportant and useful methods that are involved in this process are polymerase chain reaction "PCR#ampli_cation of DNAs\ and cloning of selected {{winner|| DNAs to obtain individual sequences[

Finally\ Chapter 6[07 covers the application of molecular cloning to chemically oriented problems[Cloning is not only important for obtaining nucleic acid sequences\ but it is also highly useful forproducing preparative quantities of well!de_ned proteins or peptides\ in natural or modi_edsequences[ The chapter not only covers general cloning strategies which are useful\ but also givesspeci_c examples where cloning has been successfully applied to chemical problems[

6[90[3[4 Topics not Covered

The topic of the chemistry of DNA is much too broad to be covered in its entirety in one volumeof text[ Indeed\ so many chemists and biochemists have worked in this _eld since the 0839s that theentire set of volumes from the Comprehensive Natural Products Chemistry series could have been_lled with reviews on DNA[ For that reason it was necessary to narrow the focus of this volume[We have attempted to choose topics most relevant to chemists who are interested in natural products[

Some notable DNA!related topics were not covered in this volume^ for example\ a comprehensivediscussion of DNA!binding proteins\ which is a very broad and widely studied topic\ was notincluded[ However\ the molecular motifs used by proteins in recognizing DNA structure are wellcovered in Chapter 6[02\ and references therein point to studies on whole proteins[ Also not discussedin the present volume is the topic of chromatin structure^ this is important because DNA in cells is\of course\ organized and protected within a multiprotein complex\ and changes in this complex areimportant for gene expression[ This topic has been reviewed[21Ð24 Higher!order structures of DNAs\such as triple helices\25Ð27 tetrastranded structures\28 and branched junctions\39 are discussed in partsof several chapters in this volume\ and have been reviewed in more detail elsewhere[ Telomerase isalso an important DNA!related topic of late^ this ribonucleoprotein helps protect natural chro!mosomes by maintaining the ends[ Good reviews on telomerase have been published[30\31 Anotherimportant class of compounds that interact with DNA are the minor groove binders[ Reviews onminor groove interactions have been written[32 Finally\ the topic of enzymatic DNA repair hasreceived much attention\ and more information on this topic is readily available[33\34 Readers whoare interested in one or more of these topics are invited to visit these other sources[

6[90[4 THE FUTURE OF DNA CHEMISTRY RESEARCH

The Editor of this volume would like to take this opportunity to speculate on where DNAresearch is heading over the coming decade[ The topics covered are far from comprehensive and arenecessarily biased toward my own views[ However\ this list represents what some chemists believe

8Overview

will be important applications of research on DNA!related topics\ and o}ers a wide array ofopportunities for new chemists entering the _eld to develop useful tools for scientists and therapiesfor medicine in the future[

6[90[4[0 Natural Products that Interact with DNA

While many natural products have already been shown to interact with DNA\ it is clear thatthere exist in nature many more natural products that remain to be identi_ed[ Many of these willalso be found to bind to DNA as well\ and some of them will no doubt be important in a medicinalsense[ While many new compounds will be found to bind in already well!de_ned motifs such asintercalation and minor groove binding\ it is highly likely that new molecular motifs for DNArecognition will be discovered through this research\ and new modes of DNA binding will beuncovered[ An example of this has been found in the interaction of chromomycin with DNA\which was found to form a surprising complex with DNA in the minor groove "Figure 6#[35 Twochromomycin molecules were found to form a co!complex with a magnesium dication[ In the future\more such novel DNA!binding modes will undoubtedly be identi_ed[ In addition\ the imaginationof chemists will continue unfettered\ and so synthetic molecules inspired by natural products willalso be shown to form novel types of complexes with DNA as well[

Figure 6 Chromomycin binds DNA by forming a dimeric complex around a magnesium ion "shown above#[35

6[90[4[1 DNA and Analogues as Diagnostic Tools

The development of DNA!based diagnostic tools is undergoing an explosion\ and there will bemany more such probes developed[ An elegant example of this are the {{molecular beacons|| ofKramer "Figure 7#\36Ð38 which are beginning to be widely applied in research[ Perhaps the greatestexpansion in this _eld will be in the practical application of many of these new methods to humanpatients[ To date\ there are few DNA!based molecular diagnostic methods which are actuallyapproved for use[ The completion of the sequencing of the human genome will make available manynew genetic targets for identi_cation[ Moreover\ the future will see the development of rapid\ high!throughput diagnostic and sequencing methodologies for individual patients[ There is clearly stillmuch chemistry to be done in this area[

6[90[4[2 DNA and Analogues as Therapeutic Agents

The concept of the use of small synthetic DNAs as potential therapeutics "antisense and antigenetherapy# dates only from the 0869s[ After years of basic research in academic labs\ the 0879s and0889s saw the undertaking of commercial development of this idea[ Since the mid!0889s there havebeen some very promising clinical results using such agents in human patients[8\09 If some of these

09 Overview

Figure 7 Structure of a ~uorescent {{molecular beacon|| for sensing the presence of complementary nucleicacids in solution[36

compounds are approved for use\ this will drive the development of an even wider number of DNAanalogues for application to an increasing number of disease states[ Many chemists will thereforeremain involved\ both in academic and industrial laboratories\ in the development of new molecularstructures and sequences of DNA analogues[ In addition\ new genetically targeted and DNA!relatedmolecular strategies will be developed for treating disease[ For example\ among the newest of thesein the literature are a number of strategies which go beyond simple DNA binding to actual genecorrection or targeted mutagenesis\49Ð41 and these will likely see increasing research over the comingyears[

6[90[4[3 DNA as a Molecular Scaffold

The regularity and increasing predictability of DNA helical structures has led increasingly to theiruse as sca}olds in the organization of other molecules or assemblies of molecules[ The developmentof a large number of convenient methods for modi_cation of DNA and conjugation of groups toDNA has made this idea increasingly feasible to carry out[ Moreover\ the ability of DNA to self!assemble in solution into large\ complex\ and stable structures makes DNA "as opposed to otherpossible sca}old!like molecules# very attractive for this purpose[ One example of this was the useof DNAs to organize gold nanoparticles^42 these were use as color!changing sensors of other geneticsequences[43 Indeed\ it seems possible "and even likely# that DNA sca}olds will also be utilized in_elds other than the biological or medical ones[44 For example\ materials research can make gooduse of a regular\ predictable sca}old for the organization of molecules on surfaces[

6[90[4[4 DNA as a Ligand

The advent of in vitro selection methods for single!stranded nucleic acids in the late 0879s45Ð47 hasled the consensus that DNA "and RNA# may be able to act as ligands for a surprisingly wide varietyof molecules[ While this may have been initially surprising because of the seeming lack of varied

00Overview

polarity and functionality in DNA\ we now know that DNA can fold to form complex shapes whichallow it to bind speci_cally in clefts of large proteins and\ in turn\ to form small clefts in itself tobind small organic molecules "see Chapter 6[06#[ Such binding can lead to biological activity "see\for example\ the published thrombin inhibitor DNAs "Figure 8#48#^ however\ there is\ of course\ noreal reason why this kind of ligand development needs to be limited to the binding of biologicallyderived molecules[ The continuing chemical development of modi_cations for DNA bases andbackbones which can be carried through in vitro selection cycles59 will make possible the tight andspeci_c binding of an even wider variety of molecules in the future[

Figure 8 Structure of a DNA molecule that binds and inhibits human thrombin[48 "a# Folded structure isshown along with "b# chemical structure of G tetrad portion of structure[

6[90[4[5 DNA as a Catalyst

The in vitro selection combinatorial strategies described above also make possible the e.cientsearching for DNAs which fold to form a catalytic site[ All that is required to search for catalysts"as opposed to simple ligands# is a means for separating catalytic molecules from inactive ones\ andseveral such strategies are under rapid development[ Although there is a lingering perception thatnatural DNA may be limited as a catalyst because of the lack of suitable functionality "see Chapter6[06#\ chemists are developing new functional groups for DNA which may well solve this potentialproblem[50\51 It seems possible and even probable\ therefore\ that synthetic chemists may in the futureemploy modi_ed DNA molecules as practical catalysts for some enantioselective transformations[

6[90[4[6 DNA as a Computer

One of the most surprising applications of DNA research has been the proposal that DNA maybe useful in computing of certain types of computationally hard problems[52 There are now a number

01 Overview

of publications describing the successful use of DNAs to solve problems that may have been solvedconsiderably more slowly if a silicon!based computer had tackled them[52\53 Although it remains tobe seen how generally useful this approach will become\ it certainly illustrates the broad applicabilityof DNA\ once considered only a natural product[

ACKNOWLEDGMENT

I thank the National Institutes of Health\ the Army Research O.ce\ the New York State O.ceof Science + Technology\ and the O.ce of Naval Research for support of various aspects of ourwork involving modi_ed DNAs[ I further acknowledge the Dreyfus Foundation for a Teacher!Scholar Award and the Alfred P[ Sloan Foundation for a Sloan Fellowship[ Finally\ I thank all myco!workers\ present and past\ for their contributions to our work[

6[90[5 REFERENCES

0[ A[ Kornberg\ Science 0858\ 052\ 0309[1[ A[ Kornberg\ in {{DNA Replication\|| Freeman\ San Francisco\ CA\ 0879\ p[ 090[2[ R[ L[ Letsinger and V[ Mahadevan\ J[ Am[ Chem[ Soc[\ 0855\ 77\ 4208[3[ S[ L[ Beaucage and M[ H[ Caruthers\ Tetrahedron Lett[\ 0870\ 11\ 0748[4[ S[ M[ Gryaznov\ D[ H[ Lloyd\ J[ K[ Chen\ R[ G[ Schultz\ L[ A[ DeDionisio\ L[ Ratmeyer\ and W[ D[ Wilson\ Proc[

Natl[ Acad[ Sci[ USA\ 0884\ 81\ 4687[5[ P[ E[ Nielsen\ M[ Egholm\ R[ Berg\ and O[ Buchardt\ Science\ 0880\ 143\ 0386[6[ A[ Van Aerschot\ I[ Verheggen\ C[ Hendrix\ and P[ Herdewijn\ An`ew[ Chem[\ Int[ Ed[ En`l[\ 0884\ 23\ 0227[7[ M[ Tarkoy\ M[ Bolli\ B[ Schweizer\ and C[ Leumann\ Helv[ Chim[ Acta\ 0882\ 65\ 370[8[ E[ Bayever\ P[ Iversen\ L[ Smith\ J[ Spinolo\ and G[ Zon\ Antisense Res[ Dev[\ 0881\ 1\ 098[

09[ W[ Roush\ Science\ 0886\ 165\ 0081[00[ D[ Kisner\ in {{01th International Round Table on Nucleosides\ Nucleotides and Their Biological Applications\|| La

Jolla\ CA\ 0885\ OP 35[01[ D[ L[ Cole\ V[ T[ Ravikumar\ A[ Krotz\ D[ C[ Capaldi\ Z[ S[ Cheruvallath\ A[ N[ Scozzari\ M[ Andrade\ and E[ F[

Gritzen\ in {{01th International Round Table on Nucleosides\ Nucleotides and Their Biological Applications\|| La Jolla\CA\ 0885\ OP 35[

02[ M[ F[ Denissenko\ A[ Pao\ M!s[ Tang\ and G[ P[ Pfeifer\ Science\ 0885\ 163\ 329[03[ M[ Hollstein\ D[ Sidransky\ B[ Vogelstein\ and C[ C[ Harris\ Science\ 0880\ 142\ 38[04[ M[ S[ Greenblatt\ W[ P[ Bennett\ M[ Hollstein\ and C[ C[ Harris\ Cancer Res[\ 0883\ 43\ 3744[05[ S[ Beck and P[ Sterk\ Curr[ Opin[ Biotechnol[\ 0887\ 8\ 005[06[ C[ M[ Fraser and R[ D[ Fleischmann\ Electrophoresis\ 0886\ 07\ 0196[07[ K[ K[ Murray\ J[ Mass Spectrom[\ 0885\ 20\ 0192[08[ M[ J[ O|Donnell!Maloney\ C[ L[ Smith\ and C[ R[ Cantor\ Trends Biotechnol[\ 0885\ 03\ 390[19[ N[ A[ Heerema\ Cancer Invest[\ 0887\ 05\ 016[10[ M[ P[ Strout and M[ A[ Caligiuri\ Curr[ Opin[ Oncol[\ 0886\ 8\ 7[11[ U[ Pindur and T[ Lemster\ Pharmazie\ 0887\ 42\ 68[12[ J[ Cummings and J[ F[ Smyth\ Ann[ Oncol[\ 0882\ 3\ 422[13[ J[ M[ Berger\ Curr[ Opin[ Struct[ Biol[\ 0887\ 7\ 15[14[ A[ D[ Bates and A[ Maxwell\ Curr[ Biol[\ 0886\ 6\ R667[15[ J[ Tazi\ F[ Rossi\ E[ Labourier\ I[ Gallouzi\ C[ Brunel\ and E[ Antoine\ J[ Mol[ Med[\ 0886\ 4\ 675[16[ M[ R[ Smith\ W[ A[ Peters III\ and C[ W[ Drescher\ Am[ J[ Obstet[ Gynecol[\ 0883\ 069\ 0566[17[ M[ A[ Kercher\ P[ Lu\ and M[ Lewis\ Curr[ Opin[ Struct[ Biol[\ 0886\ 6\ 65[18[ P[ Youderian and D[ N[ Arvidson\ Gene 0883\ 049\ 0[29[ J[ L[ Pomerantz\ C[ O[ Pabo\ and P[ A[ Sharp\ Proc[ Natl[ Acad[ Sci[ USA\ 0884\ 81\ 8641[20[ C[ Park\ J[ L[ Campbell\ and W[ A[ Goddard III\ Proc[ Natl[ Acad[ Sci[ USA\ 0881\ 78\ 8983[21[ A[ I[ Lamond and W[ C[ Earnshaw\ Science\ 0887\ 179\ 436[22[ K[ Luger and T[ J[ Richmond\ Curr[ Opin[ Struct[ Biol[\ 0887\ 7\ 22[23[ M[ Grunstein\ Nature\ 0886\ 278\ 238[24[ K[ Struhl\ Genes Dev[\ 0887\ 01\ 488[25[ G[ E[ Plum\ D[ S[ Pilch\ S[ F[ Singleton\ and K[ J[ Breslauer\ Annu[ Rev[ Biophys[ Biomol[ Struct[\ 0884\ 13\ 208[26[ J[ S[ Sun\ T[ Garestier\ and C[ He�le�ne\ Curr[ Opin[ Struct[ Biol[\ 0885\ 5\ 216[27[ L[ J[ Maher III\ Bioessays\ 0881\ 03\ 796[28[ D[ Rhodes and R[ Giraldo\ Curr[ Opin[ Struct[ Biol[\ 0884\ 4\ 200[39[ N[ C[ Seeman\ DNA Cell Biol[\ 0880\ 09\ 364[30[ C[ W[ Greider\ Proc[ Natl[ Acad[ Sci[ USA\ 0887\ 84\ 89[31[ T[ R[ Cech\ T[ M[ Nakamura\ and J[ Lingner\ Biokhimiya "Moscow#\ 0886\ 51\ 0191[32[ D[ E[ Wemmer and P[ B[ Dervan\ Curr[ Opin[ Struct[ Biol[\ 0886\ 6\ 244[33[ H[ E[ Krokan\ R[ Standal\ and G[ Slupphaug\ Biochem[ J[\ 0886\ 214\ 0[34[ S[ S[ Parikh\ C[ D[ Mol\ and J[ A[ Tainer\ Structure\ 0886\ 4\ 0432[35[ X[ L[ Gao\ P[ Mirau\ and D[ J[ Patel\ J[ Mol[ Biol[\ 0881\ 112\ 148[36[ S[ Tyagi and F[ R[ Kramer\ Nat[ Biotechnol[\ 0885\ 03\ 292[37[ G[ Leone\ H[ van Schijndel\ B[ van Gemen\ F[ R[ Kramer\ and C[ D[ Schoen\ Nucleic Acids Res[\ 0887\ 15\ 1049[

02Overview

38[ S[ Tyagi\ D[ P[ Bratu\ and F[ R[ Kramer\ Nat[ Biotechnol[\ 0887\ 05\ 38[49[ E[ S[ Belousov\ I[ A[ Afonina\ M[ A[ Podyminogin\ H[ B[ Gamper\ M[ W[ Reed\ R[ M[ Wydro\ and R[ B[ Meyer\ Nucleic

Acids Res[\ 0886\ 14\ 2339[40[ M[ Raha\ G[ Wang\ M[ M[ Seidman\ and P[ M[ Glazer\ Proc[ Natl[ Acad[ Sci[ USA\ 0885\ 82\ 1830[41[ A[ Cole!Strauss\ K[ Yoon\ Y[ Xiang\ B[ C[ Byrne\ M[ C[ Rice\ J[ Gryn\ W[ K[ Holloman\ and E[ B[ Kmiec\ Science\

0885\ 162\ 0275[42[ R[ Elghanian\ J[ J[ Storho}\ R[ C[ Mucic\ R[ L[ Letsinger\ and C[ A[ Mirkin\ Science\ 0886\ 166\ 0967[43[ C[ A[ Mirkin\ R[ L[ Letsinger\ R[ C[ Mucic\ and J[ J[ Storho}\ Nature\ 0885\ 271\ 596[44[ J[ H[ Chen and N[ C[ Seeman\ Nature\ 0880\ 249\ 520[45[ C[ Tuerk and L[ Gold\ Science\ 0889\ 138\ 494[46[ D[ L[ Robertson and G[ F[ Joyce\ Nature\ 0889\ 233\ 356[47[ A[ D[ Ellington and J[ W[ Szostak\ Nature\ 0889\ 235\ 707[48[ L[ C[ Bock\ L[ C[ Gri.n\ J[ A[ Latham\ E[ H[ Vermaas\ and J[ J[ Toole\ Nature\ 0881\ 244\ 453[59[ J[ A[ Latham\ R[ Johnson\ and J[ J[ Toole\ Nucleic Acids Res[\ 0883\ 11\ 1706[50[ J[ K[ Bashkin\ J[ K[ Gard\ and A[ S[ Modak\ J[ Or`[ Chem[\ 0889\ 44\ 4014[51[ G[ Wang and D[ E[ Bergstrom\ Tetrahedron Lett[\ 0882\ 23\ 5610[52[ L[ M[ Adleman\ Science\ 0883\ 155\ 0910[53[ F[ Guarnieri\ M[ Fliss\ and C[ Bancroft\ Science\ 0885\ 162\ 119[