Enzyme-Monitored Turnover of Escherichia coli Thioredoxin Reductase: Insights for Catalysis
| Autor: | Brett W. Lennon, Charles H. Williams |
|---|---|
| Rok vydání: | 1996 |
| Předmět: |
chemistry.chemical_classification
Thioredoxin-Disulfide Reductase Chemistry Protein subunit Thioredoxin reductase Ferredoxin-thioredoxin reductase medicine.disease_cause Biochemistry Catalysis Protein Structure Tertiary Substrate Specificity Kinetics chemistry.chemical_compound Enzyme Spectrophotometry Oxidoreductase Pyridine Escherichia coli medicine bacteria Oxidation-Reduction NADP |
| Zdroj: | Biochemistry. 35:4704-4712 |
| ISSN: | 1520-4995 0006-2960 |
| DOI: | 10.1021/bi952521i |
| Popis: | Thioredoxin reductase from Escherichia coli is a member of the pyridine nucleotide-disulfide oxidoreductase family, and contains one FAD and one redox-active disulfide per subunit. It is known that two other well-studied members of this family, lipoamide dehydrogenase and glutathione reductase, cycle between the two electron-reduced and fully oxidized forms in catalysis. Enzyme-monitored turnover shows that the spectrum of thioredoxin reductase during turnover represents fully reduced flavin with NADP(H) bound. Whether the pyridine nucleotide bound is NADPH or NADP+ is dependent on the concentration of each species, i.e., how far turnover has progressed. It is also shown that the midpoint potentials of this enzyme are increased through the differential binding of NADP+ to the oxidized and reduced form of the enzyme. When combined with other kinetic and oxidation/reduction studies of this enzyme, these results indicate that thioredoxin reductase cycles between the four-electron-reduced and two-electron-reduced forms in catalysis, and that it does so with pyridine nucleotide bound. These results clarify the mechanism of thioredoxin reductase in relation to the known structure the enzyme, and provide support for earlier work in which we proposed that this enzyme utilizes a ternary complex mechanism in catalysis. |
| Databáze: | OpenAIRE |
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