13346_Motifs and Domains in Signaling VM

8/7/2019 13346_Motifs and Domains in Signaling VM

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Motifs and Domains in Signaling

By V Mishra


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The extracellular region of different PTKs iscomposed of different domains.

For instance, the EGFR extracellular domain containstwo cysteine-rich regions. The PDGFR extracellular domain consists of five immunoglobulin-like repeats .Other domains found in the extracellular region of RPTKs include fibronectin III repeats, kringle domainsand leucine-rich motifs .The extracellular domain contains the binding sitefor RPTK ligands, which can range from soluble factors

and extracellular matrix proteins to surface proteinsexpressed on other cells.


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The extracellular domain also may be involved in the dimerization of RPTKs, aprocess critical for the activation of intrinsic tyrosine kinase activity.

The transmembrane domain in the RPTK is a hydrophobic segmentof 22 to 26 amino acids situated in the cell membrane. It is flanked by aproline-rich region in the N-terminus and a cluster of basic amino acids in theC-terminus. This combination of structures secures the transmembranedomain within the lipid bilayer. There is a low degree of homology in thetransmembrane domain, even between two closely related RPTKs, suggestingthat the primary sequence does not contain important information for signal transduction.

The cytoplasmic domain primarily consists of the catalytic domain and variousautophosphorylation sites that regulate catalytic function and serve asdocking sites for proteins that contain SH2 domains.

The protein kinase catalytic domains of RPTKs are highly conservedand similar in structure to the NRPTK catalytic domains.


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Modular Domains: Subunits of a protein mediating P-P interactionsthrough short peptides

1. The SH 1 domain:

is the catalytic domain.

The crystal structure of the Src-family tyrosine kinase hck has been solved.

Its catalytic (SH1) domain has an overall bilobal structure, with a small ATP-bindingN-terminal lobe and a large peptide-binding C-terminal lobe.

The N-lobe contains a glycine-rich sequence, GXGXXGXV , and an invariabledownstream lysine for ATP binding.

Mutation of this lysine often leads to loss of catalytic activity.

The C-lobe contains a glutamate residue which catalyzes transfer of thephosphate group from ATP to tyrosine.

The region C-terminal of the SH1 domain is called the regulatory domain because itcontains a tyrosine residue (Y527 in Src) critical for modulating kinase activity.


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2. SH2 Domain:

100 aa long, found within 115 human proteinsContains 2 alpha helices and 7 beta strandsHigh affinity for phospho tyrosine residuesRepresent largest class of pTyr recognition domainSH 2 phosphotyrosine interactions are sequence-specific for different SH 2 domains .SH 2 domains contain two binding pockets for their target peptides . O ne of these pockets

contains a positively charged arginine residue, which interacts with the negativelycharged phosphate group on the tyrosine residue .The other pocket interacts with three to five amino acids C-terminal to thephosphotyrosine and plays a major role in determining substrate specificity .However, amino acids N-terminal to the phosphotyrosine also may contributeto SH 2 domain binding .


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SH-C STRUCTURE


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In all cases examined, the arginine residue

of the SH2 domain s (FLVR)conservedsequence motif in b-strand-2 is completely buried in the hydrophobic core and formsan ion pair with the phosphate group of thePhosphotyrosine

A second arginine in the first alpha helixalso binds pTyr via an amino-aromaticinteration with the amino acid s ringstructure and a salt bridge with the

phosphate.


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Consensus sequence for ligands of various SH2 domains


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3. SH 3 Domain:

F irst identified as a conserved viral adapter protein v-Crk Protein domain of about 60aa residueO ther than adapters, proteins such as PI3K, Ras GTPase activating proteins etcalso possess themThe SH3 domain has a characteristic beta-barrel fold which consists of five or six -strands arranged as two tightly packed anti-parallel sheets .The linker regions may contain short helicesApproximately 300 SH3 domains are encoded in the human genomeThis domain is C-terminal to the unique domain and interacts withproline-rich sequences or motifs in a left-handed helix

SH3 domains mediate intermolecular protein protein interactions betweenSrc-like kinases and other proteins and intracellular interactions as well


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SH-3 CONSENSUS


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The structural domains of the Src kinase family are, in order from the N-terminus:the SH4 (Src homology 4), SH3, SH2 and SH1 domains. SH1 is the catalytic domain

SH2 and SH3 are both molecular adhesives important for protein protein interaction.SH4 resides at the very N-terminus and plays a role in membrane attachment

Between the SH4 and the SH3 domains is a region whose sequence variesconsiderably among different members of the Src family


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P leckstrin homology domain (PH domain): a protein domain of approximately120 amino acids , occurs in a wide range of proteins involved in intracellular signaling or as constituents of the cytoskeleton

This domain can bind Phosphatidylinositol lipids within biological membranes(such as Phosphatidylinositol (3,4,5)-trisphosphate and phosphatidylinositol(4,5)-bisphosphate ), and proteins such as the -subunits of heterotrimericG proteins and protein kinase C .

PH domains play a role in recruiting proteins to different membranes , thustargeting them to appropriate cellular compartment or enabling them to interactwith other components of the signal transduction pathways .

Individual PH domains possess specificities for phosphoinositides phosphorylatedat different sites within the inositol ring, e.g, some bind phosphatidylinositol

(4,5)-bisphosphate but not Phosphatidylinositol (3,4,5)-trisphosphate or Phosphatidylinositol (3,4)-bisphosphate , while others may possess therequisite affinity


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PDZ D omain :(post synaptic density protein )

Fig: Molecular structure of the PDZ domain included in the human GOPC

(Golgi-associated PDZ and coiled-coil motif-containing protein) protein

1.These domains help anchor transmembrane proteins to the cytoskeleton andhold together signaling complexes

2. Has structural domain of 80-90 amino-acids found in the signaling proteins of bacteria , yeast , plants , viruses and animals

MembraneAnchorage


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The normal cellular Abl gene product is a cytoplasmic tyrosine kinase,whereas the normal Bcr gene product is a serine/threonine kinase.However, the fused product of the Philadelphia translocation has lostimportant regulatory elements from both parent proteins and results inan oncoprotein with uncontrolled tyrosine kinase activity.

It exists as three differently sized isoforms (withMW 190, 210 or 230kDa) that reflect three different breakpoints in the Bcr gene. The largest(p230) Bcr-Abl fusion protein retains a domain from Bcr that was foundto have striking similarity with unrelated proteins such as RhoGAP andthe p85 regulatory subunit of PI-3-kinase

RhoGAP domain is a conserved segment containing 200 amino acidsconsisting of seven to nine alpha-helices connected by loops, with thecore of homology being centred on a four helix bundle that contains anarginine-containing loop thought to represent the catalytic residueneeded to complete the active site of Rho-like GTPases and/or theclosely related Rac proteins

B cr Homology ( B crH) D omain :- Summary


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Bcr-homology (BcrH)

domains Contd ..its involvement in chronic myelogenousleukaemia (CML)

A majority of patients with CML show ashortened chromosome 22 with abnormal G-

banding: the Philadelphia chromosome


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reverse translocation


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The new chromosome carries part of the genesituated at the breakpoint cluster region(Bcr) of chromosome 22 and this truncatedgene s 5 end is fused it with a 3 fragment of asecond truncated gene from chromosome 9,the A belson gene (A bl)


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the abbreviation BH was first proposed forthe Bcr homology domain

RhoGA P homology domain is another name

is a conserved segment containing 200 aminoacids consisting of seven to nine a-helices

connected by loops, with the core of homology being centred on a four helix bundle thatcontains an arginine-containing loop


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BcrH domains that lack this arginine are unableto stimulate Rho/Rac-GTPase activity but can still

bind to Rho/Rac-like proteins.

In functional RhoG A Ps, the catalytic arginine

acts like the arginine fingerit helps to orientate the catalytic Gln residue of Rho that is responsible for coordinating acatalytic water molecule


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Dbl homology (DH) domains

DH-PH pair in a protein is the signature of aguanine nucelotide exchange factor (GEF) forthe Rho family of monomeric G proteins

The PH domain is thought to directintracellular localisation to membrane boundPIP3 and the DH domain is responsible for theGEF catalytic activity


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Bcl-2 homology (BH) domains

In B-cell lymphomas, the Bcl-2 gene from thelong arm of chromosome 18 is translocatedand becomes fused with the immunoglobulinheavy chain joining region ( IgJH) from the longarm of chromosome 14.

The resulting chaemeric protein (Bcl-2-JH) isover-expressed


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Both the normal and chaemeric Blc-2 provedto be apoptosis inhibitors.

solving the Bcl-2 sequence and structure ledto the identification of a family of relatedapoptosis regulators containing conserved

Bcl-2 homology (BH) domains.

The growing Bcl-2 family includes Bax, Bad,Bcl-XL, Bak and others


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13346_Motifs and Domains in Signaling VM

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