Purification of tyrosinase to homogeneity based on its resistance to sodium dodecyl sulfate-proteinase K digestion

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Abstract

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.

Original languageEnglish (US)
Pages (from-to)122-129
Number of pages8
JournalArchives of Biochemistry and Biophysics
Volume275
Issue number1
DOIs
StatePublished - Nov 15 1989

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology

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