Purification of tyrosinase to homogeneity based on its resistance to sodium dodecyl sulfate-proteinase K digestion
| Autor: | Edward J. Yurkow, Jeffrey D. Laskin |
|---|---|
| Rok vydání: | 1989 |
| Předmět: |
Tyrosinase
Sodium Ion chromatography Biophysics Melanoma Experimental chemistry.chemical_element Biochemistry Chromatography DEAE-Cellulose Cell Line chemistry.chemical_compound Mice Animals Sodium dodecyl sulfate Amino Acids Molecular Biology chemistry.chemical_classification Gel electrophoresis Chromatography biology Chemistry Monophenol Monooxygenase Serine Endopeptidases Sodium Dodecyl Sulfate Proteinase K Molecular Weight Kinetics Enzyme Polyclonal antibodies biology.protein Electrophoresis Polyacrylamide Gel Endopeptidase K Catechol Oxidase |
| Zdroj: | Archives of biochemistry and biophysics. 275(1) |
| ISSN: | 0003-9861 |
| Popis: | A two-step method was used to purify enzymatically active tyrosinase to apparent homogeneity from B16/C3 mouse melanoma cells. The purification was based on our finding that tyrosinase was highly resistant to sodium dodecyl sulfate-proteinase K digestion. Following treatment with proteinase K, tyrosinase was separated from all detectable contaminants using DEAE-52 ion-exchange chromatography. Using this procedure, tyrosinase was purified to high specific activity and in yields greater than 50%. The purified enzyme was then used to generate high titers of specific polyclonal antibodies in rabbits. These antibodies were used to immunoprecipitate all tyrosinase isozymes from murine melanoma tumor extracts and cultured B16/C3 cells metabolically labeled with [35]methionine. |
| Databáze: | OpenAIRE |
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