SUBSTRATE-BINDING AND CONFORMATIONAL-CHANGES OF CLOSTRIDIUM-GLUTAMICUM DIAMINOPIMELATE DEHYDROGENASE REVEALED BY HYDROGEN DEUTERIUM EXCHANGE AND ELECTROSPRAY MASS-SPECTROMETRY/

C. glutamicum meso-diaminopimelate dehydrogenase is an enzyme of the L -lysine biosynthetic pathway in bacteria. The binding of NADPH and dia minopimelate to the recombinant, overexpressed enzyme has been analyze d using hydrogen/deuterium exchange and electrospray ionization/mass s pectrometry. NADPH binding reduces the extent of deuterium exchange, a s does the binding of diaminopimelate. Pepsin digestion of the deutera ted enzyme and enzyme-substrate complexes coupled with liquid chromato graphy/mass spectrometry have allowed the identification of eight pept ides whose deuterium exchange slows considerably upon the binding of t he substrates. These peptides represent regions known or thought to bi nd NADPH and diaminopimelate. One of these peptides is located at the interdomain hinge region and is proposed to be exchangeable in the ''o pen,'' catalytically inactive, conformation but nonexchangeable in the ''closed,'' catalytically active conformation formed after NADPH and diaminopimelate binding and domain closure. Furthermore, the dimerizat ion region has been localized by this method, and this study provides an example of detecting protein-protein interface regions using hydrog en/deuterium exchange and electrospray ionization.