CONFORMATIONAL FLEXIBILITY IN GLUTAMATE-DEHYDROGENASE - ROLE OF WATER IN SUBSTRATE RECOGNITION AND CATALYSIS

We have solved the structure of the binary complex of the glutamate dehydrogenase from Clostridium symbiosum with glutamate to 1.9 Å resolution. In this complex, the glutamate side-chain lies in a pocket on the enzyme surface and a key determinant of the enzymic specificity is an interaction of the substrate γ-carboxyl group with the amino group of Lys89. In the apo-enzyme, Lys113 from the catalytic domain forms an important hydrogen bond to Asn373, in the NAD+-binding domain. On glutamate binding, the side-chain of this lysine undergoes a significant movement in order to optimize its hydrogen bonding to the α-carboxyl group of the substrate. Despite this shift, the interaction between Lys113 and Asn373 is maintained by a large-scale conformational change that closes the cleft between the two domains. Modelling studies indicate that in this "closed" conformation the C-4 of the nicotinamide ring and the α-carbon atom of the amino acid substrate are poised for efficient hydride transfer. Examination of the structure has led to a proposal for the catalytic activity of the enzyme, which involves Asp165 as a general base, and an enzyme-bound water molecule, hydrogen-bonded to an uncharged lysine residue, Lys125, as an attacking nucleophile in the reaction. © 1993 Academic Press Limited.