1ju9
From Proteopedia
|
HORSE LIVER ALCOHOL DEHYDROGENASE VAL292SER MUTANT
Overview
The participation of Val-292 in catalysis by alcohol dehydrogenase and the, involvement of dynamics were investigated. Val-292 interacts with the, nicotinamide ring of the bound coenzyme and may facilitate hydride, transfer. The substitution of Val-292 with Ser (V292S) increases the, dissociation constants for the coenzymes (NAD(+) by 50-fold, NADH by, 75-fold) and the turnover numbers by 3-7-fold. The V292S enzyme, crystallized in the presence of NAD(+) and 2,3,4,5,6-pentafluorobenzyl, alcohol has an open conformation similar to the structure of the wild-type, apo-enzyme, rather than the closed conformation observed for ternary, complexes with wild-type enzyme. The V292S substitution perturbs the, conformational equilibrium of the enzyme and decreases the kinetic, complexity, which permits study of the hydride transfer step with, steady-state kinetics. Eyring plots show that the DeltaH for the oxidation, (V(1)) of the protio and deuterio benzyl alcohols is 13 kcal/mol and that, the kinetic isotope effect of 4.1 is essentially temperature-independent., Eyring plots for the catalytic efficiency for reduction of benzaldehyde, (V(2)/K(p)) with NADH or NADD are distinctly convex, being, temperature-dependent from 5 to 25 degrees C and temperature-independent, from 25 to 50 degrees C; the kinetic isotope effect of 3.2 for V(2)/K(p), is essentially independent of the temperature. The temperature, dependencies and isotope effects for V(1) and V(2)/K(p) are not adequately, explained by semiclassical transition state theory and are better, explained by hydride transfer occurring through vibrationally assisted, tunneling.
About this Structure
1JU9 is a Single protein structure of sequence from Equus caballus with ZN and NAD as ligands. Active as Alcohol dehydrogenase, with EC number 1.1.1.1 Full crystallographic information is available from OCA.
Reference
Contributions of valine-292 in the nicotinamide binding site of liver alcohol dehydrogenase and dynamics to catalysis., Rubach JK, Ramaswamy S, Plapp BV, Biochemistry. 2001 Oct 23;40(42):12686-94. PMID:11601993
Page seeded by OCA on Tue Nov 20 18:34:58 2007
