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Kinetics of Protein-Protein Interactions

Kinetics of Protein-Protein Interactions. November 2002. Content. Preview – Basic kinetics Protein-protein Kinetics – Basic view Electrostatic steering – study review. 1.Preview – Basic kinetics. Reaction Rate (V) – Change of concentration over time. Basic Reaction

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Kinetics of Protein-Protein Interactions

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  1. Kinetics of Protein-ProteinInteractions November 2002

  2. Content • Preview – Basic kinetics • Protein-protein Kinetics – Basic view • Electrostatic steering – study review

  3. 1.Preview – Basic kinetics Reaction Rate (V)– Change of concentration over time • Basic Reaction A C Rate slows as concentration of A decreases

  4. Reaction Rate constant • Example 1: H2 + F2 2HI V = K [F2] [H2] • Example 2: F2 + 2NO2 2NO2F V = K [F2] [NO2] • A + B C V = K [A] [B] Rate is dependant on preliminary concentration of reactants Rate (at first stage of reaction) Kinetic Constant Would expect [NO2][NO2] K1 • It appears things are not that • simple: • Mechanism • Different K’s (K1 – Slow) F2 + NO2 NO2F + F Fast Equilibrium NO2 + F NO2F

  5. Factors Influencing K • Arrhenius Activation Energy – Limiting Barrier • Constant: • Size • Orientation • Solvent • electrostatics

  6. 2.Protein-Protein Kinetics – Basic View A B A B • Kd= Kdissociation / Kassociation (dissociation=off, association=on) • ΔG = -RTln(Kd) • Physiological 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 A B B A 1000 – 2000A A B A B

  7. A more elaborate representation A - A A A + A A B B B - + B B B Transition-State Diffusion + Possible Steering Intermediate Desolvation, VDW, Electrostatics √ ? ? √ ? √ Random Diffusion Electrostatic Steering Encounter Complex Transition Intermediate Final Complex

  8. Reaching the Encounter Complex A - + - + B B A A B √ ? ? Random Diffusion Electrostatic Steering Encounter Complex • Random 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 Attraction Steering • Energetic factors: Electrostatic ∆S

  9. An example of electrostatic steering

  10. Barnase-Barstar Electrostatic potential Landscape

  11. 3.Evaluation of steering effect (Camacho, Vajda) Complex separation (5A) + XY rotation • 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).

  12. Evaluation of steering effect (Wade) • ccp:cc - cytochrome c peroxidase:cytochrome c • ache:fas - acetylcholinesterase: fasciculin-2 • Bn:bs - Barnase-Barstar • hyhel5:hel - HyHEL-5 antibody: hen egg white lysozyme; • hyhel10:hel - HyHEL-10 antibody:hen egg white lysozyme

  13. Evaluation of steering effect (Wade)

  14. Evaluation of steering effect (Wade)

  15. Structure – Tem1 βLactamase

  16. Structure – Tem1 β Lactamase

  17. Structure – BLIP-ΙΙ

  18. Bound – Blip-ΙΙ & TEM1

  19. Bound – Blip-ΙΙ & TEM1

  20. Mutations on BLIP outside the active site

  21. Results

  22. Results

  23. Possible Transition state orientation • Still water molecules awaiting extraction • Possibly a core of atoms in proximity with final orientation

  24. Bound Model Camacho/wade – Electrostatic minima Encounter complex modeling Barnase - Barstar

  25. Bound Model Janin – 50% surface area + rotational limit Encounter complex modeling Barnase - Barstar

  26. Bound Model Vijayakumar – solvent separation + (2 angles – 3dg limit) Encounter complex modeling Barnase - Barstar

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