PROTEIN STRUCTURE AT ACTION: BIND  TRANSFORM  RELEASE PROTEIN PHYSICS LECTURE 24-25 PROTEIN STRUCTURE AT ACTION: BIND  TRANSFORM  RELEASE

BIND: repressors - turn - 

Zn- fingers DNA & RNA BINDING Leu-zipper

BIND   RELEASE: REPRESSOR -BINDING-INDUCED DEFORMATION MAKES REPRESSOR ACTIVE, and IT BINDS TO DNA

Immunoglobulin

Standard positions of active sites in protein folds

There are some with catalytic (Ser-protease) site

Preferential binding of TS: RIGID enzyme BIND  TRANSFORM  RELEASE Catalysis: stabilization of the transition state (TS) Theory: Pauling & Holden Preferential binding of TS: RIGID enzyme

Catalysis: stabilization of the transition state (TS) Theory: Pauling & Holden Experimental verification: Fersht reputed TS __________ ______ P

/ / / / Catalysis: stabilization of the transition state (TS) Theory: Pauling & Holden Experimental verification: Fersht / This protein engineering reduces the rate by 1000000 / / reputed TS / __________ ______ P Preferential binding of TS: RIGID enzyme

Catalytic antibodies ABZYM = AntyBody enZYM Transition state (TS) BIND  TRANSFORM  RELEASE Catalytic antibodies ABZYM = AntyBody enZYM Transition state (TS) Preferential binding of TS: RIGID enzyme Antibodies are selected to TS-like molecule

BIND  TRANSFORM  RELEASE: ENZYME chymotrypsin Note: small active site

Different folds with the same active site: Sometimes: Different folds with the same active site: the same biochemical function

 non-active “cat. site” active cat. site POST-TRANSLATIONAL MODIFICATION Sometimes, only the CHAIN CUT-INDUCED DEFORMATION MAKES THE ENZYME ACTIVE READY  non-active “cat. site” active cat. site CUT Chymotripsinogen Chymotripsin

Chymotrypsin catalyses hydrolysis of a peptide Spontaneous hydrolysis: very slow

SER-protease: catalysis

CHYMOTRYPSIN ACTIVE SITE with INHIBITOR

Preferential binding of TS: RIGID enzyme F = k1x1 = - k2x2 Ei = (ki /2)(xi)2 = F2/(2ki ) Hooke’s & 2-nd Newton’s Energy is concentrated laws in the softer body. Effective catalysis: when substrate is softer than protein Kinetic energy cannot be stored for catalysis Friction stops a molecule within picoseconds: m(dv/dt) = -(3D)v [Stokes law] D – diameter; m ~ D3 – mass;  – viscosity tkinet  10-13 sec  (D/nm)2 in water

PROTEIN STRUCTURE AT ACTION: BIND  TRANSFORM  RELEASE RIGID CATALITIC SITE INDEPENDENT ON OVERALL CHAIN FOLD

MOTIONS

from one active site to another Double sieve: movement of substrate from one active site to another  tRNAIle

Movement in two-domain enzyme: One conformation for binding (and release), another for catalysis

Two-domain dehydrogenases: Universal NAD-binding domain; Individual substrate-binding domain

Movement in quaternary structure: Hemoglobin vs. myoglobin

Механохимический цикл Миозин Актин АТФ  АДФ + Ф 15 ккал/моль в клеточных условиях Механохимический цикл

Mechanochemical cycle Myosin Actin Mechanochemical cycle

SUMMARY

PROTEIN PHYSICS Interactions Structures Selection States & transitions

Intermediates & nuclei Structure prediction & bioinformatics Protein engineering & design Functioning