involvement of essential lysine residues in the catalytic activity of glucose 6-phosphate dehyrogenase purified from streptomyces aureofaciens

glucose 6- phosphate dehydrogenase from streptomyces aureofaciens was purified andinactivated by pyridoxal 5′-phosphate (plp). the inactivation was a pseudo-first order and time-dependentreaction. complete inactivation was achieved at 0.2mm plp within 16 minutes. the type of inhibition wascompetitive with respect to glucose 6- phosphate. spectral characteristics of plp-enzyme complexcorresponded to the formation of a schiff’s base between plp and lysine residue(s) of the enzyme. intrinsicprotein fluorescence sharply decreased upon plp modification with about a 10 nm red shift. the presence ofglucose 6-phosphate in the incubation mixture prevented the fluorescence change. fluorescence studiesrevealed that nad+ and nadp+ binding induces different conformational changes in pyridoxylated enzyme.the stochiometry of plp binding to the enzyme showed that 2 moles of lysine residues were modified permole of enzyme. the data indicated that the modified lysine residues are involved in substrate binding and/orcatalytic activity of this enzyme.