Detection and purification of glucose 6-phosphate dehydrogenase, malic enzyme, and NADP-dependent isocitrate dehydrogenase by blue native polyacrylamide gel electrophoresis
| Autor: | Jeff Middaugh, Ranji Singh, Vasu D. Appanna, Ryan J. Mailloux, Daniel Chenier, Robin Beriault |
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| Rok vydání: | 2005 |
| Předmět: |
Chromatography
Clinical Biochemistry Malic enzyme Native Polyacrylamide Gel Electrophoresis Dehydrogenase Gel electrophoresis of proteins Biology Glucosephosphate Dehydrogenase Pseudomonas fluorescens Biochemistry Isocitrate Dehydrogenase Analytical Chemistry chemistry.chemical_compound Isocitrate dehydrogenase chemistry Malate Dehydrogenase Glucose-6-phosphate dehydrogenase Electrophoresis Polyacrylamide Gel Formazan Polyacrylamide gel electrophoresis |
| Zdroj: | Electrophoresis. 26(15) |
| ISSN: | 0173-0835 |
| Popis: | We describe a blue native polyacrylamide gel electrophoretic technique that allows the facile detection, quantitation and purification of three NADPH-producing enzymes. Glucose 6-phosphate dehydrogenase, malic enzyme and NADP-dependent isocitrate dehydrogenase were detected simultaneously. Activity staining based on the formation of NADPH from the respective substrates and the subsequent precipitation of formazan enabled the relative quantitation of enzymatic activities, while Coomassie staining on one-dimensional or two-dimensional gels helped monitor the amount of protein associated with these enzymatic activities. This technique provides a simple and effective route to obtain homogeneous protein for further analyses and also enables the screening of these NADPH-producing enzymes in various cellular systems. |
| Databáze: | OpenAIRE |
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