Isocitrate dehydrogenase from Rhodopseudomonas spheroides: Kinetic mechanism and further characterization

Product inhibition studies with Rhodopseudomonas spheriodes NADP + specific isocitrate dehydrogenase indicate that the enzyme mechanism involves the ordered addition of the substrates NADP + and threo- d s -isocitrate and the ordered release of products CO 2 (HCO s − ), 2-ketoglutarate, and NADPH. In addition, the presence of a ternary complex consisting of enzyme, NADP + , and 2-ketoglutarate is indicated. Binding studies with radioactive substrates support the kinetically derived mechanism. The Rhodopseudomonas enzyme is dimeric and contains but a single active site. Different combinations of substrate were ineffective in causing gross changes in molecular structure as monitored by gel filtration techniques. A comparison of the amino acid composition of this enzyme with the bacterial enzyme from Azotobacter vinelandii indicate very significant differences in the amino acid compositions.