The effect of Val22 on the catalytic activity of mandelate racemase from Pseudomonas putida

Abstract:

Located in the 20s loop of mandelate racemase from Pseudomonas putida, Val22 is one of the amino acids related with substrate binding. In this study, a 75.9% decrease of activity was observed after Val was substituted by Arg. In addition to the reduction of hydrophobic interaction between enzyme and substrate, electrostatic repulsion could also lead to the decrease of activity. Molecular dynamic was utilized and showed that the electrostatic potential of substrate and amino acid of No. 22 increased from 0.036 kJ/mol to 0.124 kJ/mol. This result indicated that the increased activity in the mutant was mainly attributed to the shift of side chain polarity and the electrostatic repulsion between side chain and substrate. An increase of 283 kJ/mol in the potential energy of system suggested that the changes of side chain polarity and electric property brought about the increase of potential energy in binding state and the dramatical decline of activity. Therefore hydrophobic interaction and electrostatic interaction should be considered with steric bulk when rationally designing the binding pocket of mandelate racemase.