Purification and properties of 8-hydroxy-5-deazaflavin-dependent NADP+ reductase from Methanococcus vannielii.

Abstract

The 8-hydroxy-5-deazaflavin-dependent NADP+ reductase component of the formate NADP+ oxidoreductase system of Methanococcus vannielii has been purified to homogeneity. The enzyme is specific for NADP+ and 8-hydroxy-5-deazaflavin. It catalyzes the reaction: 1,5-Dihydro-8-hydroxy-5-deazaflavin anion + NADP+ in equilibrium 8-hydroxy-5-deazaflavin + NADPH. The apparent molecular weight of the native enzyme is 85,000. A subunit molecular weight of 43,000 determined by sodium dodecyl sulfate gel electrophoresis indicates that the native enzyme is a dimer. The optimal temperature for catalytic activity is 17-20 degrees C and the pH maxima are 7.9 and 4.8 for the forward and reverse reactions, respectively. The kcat value of the forward reaction is 24 times greater than that of the reverse reaction, thus the production of NADPH at pH 7.0 is more favorable than its consumption. The reductase contains one or more sulfhydryl groups which are essential for catalytic activity.

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@article{Yamazaki1980PurificationAP, title={Purification and properties of 8-hydroxy-5-deazaflavin-dependent NADP+ reductase from Methanococcus vannielii.}, author={Soh Yamazaki and Luong Tsai}, journal={The Journal of biological chemistry}, year={1980}, volume={255 13}, pages={6462-5} }