Purification and Characterization ofζ-Crystallin from the Camel Lens
| Autor: | Nayyar Rabbani, Abdullah S. Alhomida, Ali S. Duhaiman, Abdulaziz A. Al-Jafari |
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| Rok vydání: | 1995 |
| Předmět: |
Camelus
Size-exclusion chromatography Biophysics Quinone oxidoreductase Biochemistry Chromatography Affinity Sepharose Affinity chromatography Crystallin Lens Crystalline Animals Molecular Biology Polyacrylamide gel electrophoresis Chromatography biology Chemistry Cell Biology Hydrogen-Ion Concentration Phenanthrenes Crystallins Enzyme assay Molecular Weight Kinetics Isoelectric point Chromatography Gel biology.protein Thermodynamics Electrophoresis Polyacrylamide Gel Isoelectric Focusing Oxidation-Reduction NADP |
| Zdroj: | Biochemical and Biophysical Research Communications. 215:632-640 |
| ISSN: | 0006-291X |
| DOI: | 10.1006/bbrc.1995.2511 |
| Popis: | zeta-crystallin a novel NADPH: quinone oxidoreductase was purified from the cortex of the camel (Camelus dromedarius) lens to homogeneity by Sepharose CL-6B gel filtration column and 2', 5' ADP-Sepharose 4B affinity column chromatography in the presence of dithiothreitol. The purified zeta-crystallin has a molecular weight of 140 kDa, as determined by Superose 12 gel filtration column. SDS-PAGE showed a single polypeptide band of molecular weight 35 kDa, suggesting that the native enzyme is composed of four identical subunits. The isoelectric point of the enzyme was 7.6 on native polyacrylamide gel. The enzyme was purified 20-fold over homogenate with a specific activity of 26.0 Units/mg protein, and an overall recovery of 53%. This enzyme was NADPH specific and followed Michaelis-Menten kinetics. Km values for the reduction of 9,10-phenanthroquinone and oxidation of NADPH were 17 microM and 6.9 microM, respectively, at pH 7.8. The Vmax values of the enzyme for 9,10 phenanthroquinone and NADPH were 32 mumole min-1, mg-1 protein and 22.7 mumole min-1 mg-1 protein, respectively. |
| Databáze: | OpenAIRE |
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