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Biotinyl-Glucose-6-Phosphate Dehydrogenase Preparation, Kinetics, and Modulation by Avidin

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Abstract

The kinetics of free glucose-6-phosphate dehydrogenase (G-6-PDH), biotinylated G-6-PDH, and biotinylated G-6-PDH complexed with avidin were investigated. The kinetics of the free enzyme were consistent with a sequential rather than a ping-pong mechanism. The kinetics of the biotinylated enzyme were similar to that of the free enzyme, but the kinetic constants were different; theK m value for NADP was halved, whereas theK m for G-6-P decreased only slightly. In the presence of avidin, theK m of biotinylated G-6-PDH for G-6-P nearly doubled whereas theK m for NADP did not change significantly. Avidin complexed with biotinylated G-6-PDH inhibited the enzyme from acting. Based upon these reactions, it was possible to devise assays for either free biotin or free avidin using biotinylated G-6-PDH as the indicator enzyme. Concentrations of biotin between 40 and 60 mg/mL, or of 25–95 Μg/mL of avidin could be measured within 2 min through the use of biotinylated G-6-PDH.

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Authors and Affiliations

  1. Department of Developmental and Cell Biology, University of California, 92717, Irvine, California, USA

    That T. Ngo, Howard M. Lenhoff & Jeanne Ivy

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  1. That T. Ngo
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  2. Howard M. Lenhoff
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  3. Jeanne Ivy
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Ngo, T.T., Lenhoff, H.M. & Ivy, J. Biotinyl-Glucose-6-Phosphate Dehydrogenase Preparation, Kinetics, and Modulation by Avidin. Appl Biochem Biotechnol 7, 443–454 (1982). https://doi.org/10.1007/BF02799175

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  • DOI: https://doi.org/10.1007/BF02799175

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