kinetics of protein-protein interactions november 2002
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Kinetics of Protein-ProteinKinetics of Protein-ProteinInteractionsInteractions
November 2002November 2002
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ContentContent
Preview – Basic kineticsPreview – Basic kinetics Protein-protein Kinetics – Basic viewProtein-protein Kinetics – Basic view Electrostatic steering – study reviewElectrostatic steering – study review
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1.Preview – Basic kinetics1.Preview – Basic kinetics
Reaction Rate (V) – Change of concentration over timeReaction Rate (V) – Change of concentration over time
Basic Reaction Basic Reaction
A CA C
Rate slows asRate slows as concentration concentration of A decreasesof A decreases
Rate slows asRate slows as concentration concentration of A decreasesof A decreases
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Reaction Rate constant Reaction Rate constant
A + B CA + B C
V = K [A] [B]V = K [A] [B]
Rate (at first Rate (at first stage stage
of reaction)of reaction)
Rate (at first Rate (at first stage stage
of reaction)of reaction)KineticKinetic
ConstantConstant
KineticKinetic ConstantConstant
Rate is dependant onRate is dependant on preliminary concentration of reactantspreliminary concentration of reactants
Rate is dependant onRate is dependant on preliminary concentration of reactantspreliminary concentration of reactants
Example 1: Example 1:
HH22 + F + F2 2 2HI 2HI
V = K [FV = K [F22] [H] [H22]]
(K1 – Slow)(K1 – Slow) F F22 + NO + NO2 2 NO NO22F + FF + F
Fast EquilibriumFast Equilibrium NO NO22 + F NO + F NO22FF
Would expectWould expect [NO[NO22][NO][NO22]]
Would expectWould expect [NO[NO22][NO][NO22]]
It appears things are not thatIt appears things are not thatsimple: simple: •MechanismMechanism•Different K’sDifferent K’s
It appears things are not thatIt appears things are not thatsimple: simple: •MechanismMechanism•Different K’sDifferent K’s
K1
Example 2:Example 2:
FF22 + 2NO + 2NO2 2 2NO 2NO22FF
V = K [FV = K [F22] [NO] [NO22]]
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KT
Ea
AeK
Constant:Constant:•SizeSize•OrientationOrientation•SolventSolvent•electrostaticselectrostatics
Constant:Constant:•SizeSize•OrientationOrientation•SolventSolvent•electrostaticselectrostatics
Activation Energy –Activation Energy –Limiting BarrierLimiting Barrier
Activation Energy –Activation Energy –Limiting BarrierLimiting Barrier
ArrheniusArrhenius
Factors Influencing KFactors Influencing K
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2.Protein-Protein Kinetics – Basic View2.Protein-Protein Kinetics – Basic View
Kd= Kdissociation / Kassociation (dissociation=off, association=on)Kd= Kdissociation / Kassociation (dissociation=off, association=on) ΔΔG = -RTln(Kd)G = -RTln(Kd)
ABKoff
KonBA
A B A B
Physiological conditions Physiological conditions Possible concentration of a unique Protein in a cell Possible concentration of a unique Protein in a cell
10^-6 – 10^-8 M 10^-6 – 10^-8 M Protein diameter 50 – 100 A (Protein surface ~8,000 Protein diameter 50 – 100 A (Protein surface ~8,000
A)A) Free Walk collision with interacting designated Free Walk collision with interacting designated
protein ~ 1000A – 2000A protein ~ 1000A – 2000A
A
1000 – 2000A
A
A
A
B
B
B
B
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A more elaborate representationA more elaborate representation
Diffusion + PossibleSteering
Desolvation, VDW, Electrostatics
Intermediate
Transition-State
RandomDiffusion
ElectrostaticSteering
EncounterComplex
Final Complex
Transition
A
B
A
+ -
B+-
A
B
AB
AB
AB
Intermediate? ? ?√ √ √
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Reaching the Encounter ComplexReaching the Encounter Complex
Random diffusion according to the Random diffusion according to the Smolochowski-Einstein equation - ~ Smolochowski-Einstein equation - ~ 10^9 - 10^10 1/MS10^9 - 10^10 1/MS
With geometrical constraints - ~ With geometrical constraints - ~ 10^5 – 10^6 1/MS10^5 – 10^6 1/MS
Adding electrostatic steering could Adding electrostatic steering could enhance rate to 10^9 1/MS enhance rate to 10^9 1/MS
A
B
A
+ -
B+-
A
B
RandomDiffusion
ElectrostaticSteering
EncounterComplex
? ?√
Energetic factors:Energetic factors:
∆S
Electrostatic
AttractionAttraction AttractionAttraction SteeringSteeringSteeringSteering
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An example of electrostatic steeringAn example of electrostatic steering
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Barnase-Barstar Electrostatic potential LandscapeBarnase-Barstar Electrostatic potential Landscape
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3.Evaluation of steering effect (Camacho, Vajda)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 0.15 M and protein dielectric 4; C. kallikrein A and pancreatic trypsin inhibitor (2KAI), ionic strength 0.15 M and protein dielectric 4;D. barnase and barstar (1BGS); E. subtilisin and chymotrypsin inhibitor (2SNI); F. subtilisin and eglin-c (1CSE), ionic strength 0.15 M and protein dielectric 4;G. trypsin and bovine pancreatic trypsin inhibitor (2PTC).
A. Chymotrypsin with turkey ovomucoid third domain (1CHO); B. human leukocyte elastase with turkey ovomucoid third domain (1PPF), ionic strength 0.15 M and protein dielectric 4; C. kallikrein A and pancreatic trypsin inhibitor (2KAI), ionic strength 0.15 M and protein dielectric 4;D. barnase and barstar (1BGS); E. subtilisin and chymotrypsin inhibitor (2SNI); F. subtilisin and eglin-c (1CSE), ionic strength 0.15 M and protein dielectric 4;G. trypsin and bovine pancreatic trypsin inhibitor (2PTC).
Complex separation (5A) + XY rotationComplex separation (5A) + XY rotation
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Evaluation of steering effect (Wade)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
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
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Evaluation of steering effect (Wade)Evaluation of steering effect (Wade)
N
i
kTxEN
x1
12)/),(exp(
1)(
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Evaluation of steering effect (Wade)Evaluation of steering effect (Wade)
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Structure – Tem1 Structure – Tem1 ββ LactamaseLactamase
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Structure – Tem1 Structure – Tem1 ββ Lactamase Lactamase
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Structure – BLIP-Structure – BLIP-ΙΙΙΙ
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Bound – Blip-Bound – Blip-ΙΙΙΙ & TEM1 & TEM1
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Bound – Blip-Bound – Blip-ΙΙΙΙ & TEM1 & TEM1
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Mutations on BLIP outside the active siteMutations on BLIP outside the active site
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ResultsResults
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ResultsResults
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Possible Transition state orientationPossible Transition state orientation
Still water molecules awaiting extraction Still water molecules awaiting extraction Possibly a core of atoms in proximity with final orientation Possibly a core of atoms in proximity with final orientation
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Encounter complex modelingEncounter complex modeling
Bound ModelBound Model Camacho/wade – Electrostatic minimaCamacho/wade – Electrostatic minima
Barnase - BarstarBarnase - Barstar
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Encounter complex modelingEncounter complex modeling
Bound ModelBound Model Janin – 50% surface area + rotational Janin – 50% surface area + rotational limitlimit
Barnase - BarstarBarnase - Barstar
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Encounter complex modelingEncounter complex modeling
Bound ModelBound Model Vijayakumar – solvent separation + (2 Vijayakumar – solvent separation + (2 angles – 3dg limit)angles – 3dg limit)
Barnase - BarstarBarnase - Barstar