lecture 8

Question 1

functions for interaction domains

Answer 1

protein localising and trafficking
recognition of spefici sites on port translational mods such a phosphorylation, acetylation, methylation hyroxylation and ubiquitination
adaptors that link distinct proteins into a common pathway
orderly assembly and disassembly of multi-protein complexes
regulating the catalytic activity and substrate specificity of enzymatic domains

Question 2

what is the dynamic control of cell signalling networks exerted by

Answer 2

intrinsic changes in protein conformations that affect specificity and catalytic rates at active sites
changes in specific binding via interaction domains

Question 3

oncogenis transformation by v-Src (from rous sarcoma virus)

Answer 3

c-src has 2 modes of intrinsic inhibition by interactions between -
sh2 domain and phosphorylated Y527
sh3 domain and proline region

v-src lacks c terminal tail containing the inhibitory phosphoY527
kinase is active as a tyrosine kinase
sh2 interacts with pY on targets
sh3 interacts with proline on targets

Question 4

SH2 domain and the prototypical phosphorecognition module

Answer 4

100 amino acid module with central beta sheet of 4-6 beta strands and 2 alpha helices found in approx 120 human proteins
binding site lies across the sheet structure flanked by 2 helices
conventional sh2 domains have a conserved deep pocket domain that recognises pY and a more variable specificity pocket that binds 3-6 residues c terminal to pY
sh2 proteins localise to tyrosine phosphorylated sites
function of sh2 is to specifically recognise phosphorylated state of tyrosine residues
contain sh2 containing intracellular signalling proteins can bind to activated PDGT receptors

Question 5

general properties of interaction domains

Answer 5

typically are independently folding of 35-150 amino acids
interaction domains lack intrinsic enzymatic activity
serve as linkers between signalling components
retain their binding properties when expressed independently
have juxtaposed n and c terminals that allows easy insertion into a loop on a preexisting protein
interaction domains have been selected during evolution for their adaptibility

Question 6

examples of modular interaction domains in cellular regulation

Answer 6

70% of human proteins have a modular architecture
PTB domains bind phosphtyrosine - containing peptides with a core NPXpY motif and PH domain which often binds specific phospholipids have the same fold
sh3 domains bind to proline rich motifs that adopts a PP11 helix
PDZ domain binds to a short region of c terminus of other specific proteins

Question 7

functions of interaction domains

Answer 7

determine the subcellular localisation and trafficking of proteins
mediate docking onto specific sites of post translational mods
adapters that link distinct proteins into a common pathway
orderly assembly and disassembly of multiprotein complexes
directing the substrate specificity of enzymatic domains

Question 8

protein localisation and trafficking

Answer 8

when p1-4,5-p2 is converted to p1-3,4,5-p3 by p1-3kinase, PKB/Akt is recruited to the plasma membrane via its P1P3 pleckstrin homology domain interaction where it can be phosphorylated by PDK1

Question 9

how does the generation of a phosphomotif by phosphorylation serve to define networks of interacting proteins

Answer 9

phosphomotif may by recognised by a protein interaction module on another protein 
recruit and confine signalling proteins to an appropriate subcellular location
determine specificity with which enzymes interact with their targets

Question 10

how do different xbox proteins in CRLs recognise specific motifs in different target substrates

Answer 10

due to their different binding domains

Question 11

adaptors may link distinct proteins into a common complex pathway

Answer 11

depending which pathways are active, adaptor proteins link different transcription factor complexes to gene promoters
adaptor proteins direct the flow of information from activated receptors into intracellular targets
the flow of info within the cell is regulated and integrated by combinational use of adaptor protein domains that recognise specific ligands

Question 12

therapeutic possibilities for protein:protein interactions (stability)

Answer 12

fungal phytotoxin fusicoccin stabilises interactions between c terminal 14-3-3-binding phosphate of plant plasma membrane H+ - ATPase and 14-3-3 proteins, thus leading to permanent activation of the proton pump and stromatal pores which causes wilting
fusicoccin is being tested on humans

Question 13

therapeutic possibilities for protein:protein interactions (induce)

Answer 13

the immunosuppressant rapamycin, a small cell permeable molecule mediates protein dimerisation between FKBP12 and FRB domain of mTOR

Question 14

therapeutic possibilities for protein:protein interactions (block)

Answer 14

SH2 domains mediate numerous cancer promoting protein-protein complexes to block specific sh2 phospho-protein interactions
drugs are being developed to bind to sh2 preventing binding of phosophoproteins and drugs that mimic sh2 domain and bing phophorylated ligands