Kinetics of Protein-ProteinInteractionsNovember 2002

ContentPreview Basic kineticsProtein-protein Kinetics Basic viewElectrostatic steering study review

1.Preview Basic kineticsReaction Rate (V) Change of concentration over time Basic Reaction

A CRate slows as concentration of A decreases

Reaction Rate constant A + B C

V = K [A] [B]Rate (at first stage of reaction)Kinetic ConstantRate is dependant on preliminary concentration of reactants(K1 Slow) F2 + NO2 NO2F + FFast Equilibrium NO2 + F NO2F

Would expect [NO2][NO2]It appears things are not thatsimple: MechanismDifferent KsK1

Constant:SizeOrientationSolventelectrostaticsActivation Energy Limiting BarrierArrhenius

Factors Influencing K

2.Protein-Protein Kinetics Basic ViewKd= Kdissociation / Kassociation (dissociation=off, association=on)G = -RTln(Kd)ABABPhysiological conditions Possible concentration of a unique Protein in a cell 10^-6 10^-8 M Protein diameter 50 100 A (Protein surface ~8,000 A)Free Walk collision with interacting designated protein ~ 1000A 2000A

A1000 2000AAAABBBB

A more elaborate representationDiffusion + PossibleSteeringDesolvation, VDW, ElectrostaticsIntermediateTransition-StateRandomDiffusionElectrostaticSteeringEncounterComplexFinal ComplexTransitionIntermediate???

Reaching the Encounter ComplexRandom diffusion according to the Smolochowski-Einstein equation - ~ 10^9 - 10^10 1/MS

With geometrical constraints - ~ 10^5 10^6 1/MS

Adding electrostatic steering could enhance rate to 10^9 1/MS Energetic factors:SElectrostaticAttraction Steering

An example of electrostatic steering

Barnase-Barstar Electrostatic potential Landscape

3.Evaluation of steering effect (Camacho, Vajda)

A. Chymotrypsin with turkey ovomucoid third domain (1CHO); B. human leukocyte elastase with turkey ovomucoid third domain (1PPF), ionic strength 0.15M and protein dielectric 4; C. kallikrein A and pancreatic trypsin inhibitor (2KAI), ionic strength 0.15M and protein dielectric 4;D. barnase and barstar (1BGS); E. subtilisin and chymotrypsin inhibitor (2SNI); F. subtilisin and eglin-c (1CSE), ionic strength 0.15M and protein dielectric 4;G. trypsin and bovine pancreatic trypsin inhibitor (2PTC).

Complex separation (5A) + XY rotation

Evaluation of steering effect (Wade)

ccp:cc - cytochrome c peroxidase:cytochrome cache:fas - acetylcholinesterase: fasciculin-2Bn:bs - Barnase-Barstarhyhel5:hel - HyHEL-5 antibody: hen egg white lysozyme;hyhel10:hel - HyHEL-10 antibody:hen egg white lysozyme

Evaluation of steering effect (Wade)

Evaluation of steering effect (Wade)

Structure Tem1 Lactamase

Structure Tem1 Lactamase

Structure BLIP-

Bound Blip- & TEM1

Bound Blip- & TEM1

Mutations on BLIP outside the active site

Results

Results

Possible Transition state orientationStill water molecules awaiting extraction Possibly a core of atoms in proximity with final orientation

Encounter complex modelingBound ModelCamacho/wade Electrostatic minimaBarnase - Barstar

Encounter complex modelingBound ModelJanin 50% surface area + rotational limitBarnase - Barstar

Encounter complex modelingBound ModelVijayakumar solvent separation + (2 angles 3dg limit)Barnase - Barstar