Isocitrate dehydrogenase from Rhodopseudomonas spheroides: kinetic mechanism and further characterization |
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Authors: | B E Buzdygon J E Braginski A E Chung |
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Institution: | Department of Biochemistry, University of Pittsburgh, Pittsburgh, Pennsylvania 15261 U.S.A. |
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Abstract: | 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-ds-isocitrate and the ordered release of products CO2 (HCOs?), 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. |
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