Rabbit P450 2E1 expressed in CHO-K1 cells has a short half-life

Suzanne Barmada, Eric Kienle, Dennis R. Koop

Research output: Contribution to journalArticle

27 Scopus citations

Abstract

Rabbit P450 2E1 was stably expressed in Chinese hamster ovary cells after cotransfection with pRC/CMV-2E1 and pFR400 which expresses murine dihydrofolate reductase with a single arginine to leucine substitution at position 22. This mutation permits amplification of expression with increasing methotrexate concentrations in CHO-K1 cells that are not dihydrofolate reductase deficient. After amplification with 1 μM methotrexate, a representative clone expressed about 15 pmol of P450 2E1/mg microsomal protein. Cells from a single 35-mm plate catalyzed the formation of 1.02 nmol 6-hydroxychlorzoxazone/106 cells/h or about 127 pmol/mg total cell protein/min. The enzyme was rapidly labeled when pulsed with [35S]-methionine. Initial pulse-chase experiments indicate that the expressed protein has a half-life of 4.8 h.

Original languageEnglish (US)
Pages (from-to)601-607
Number of pages7
JournalBiochemical and Biophysical Research Communications
Volume206
Issue number2
DOIs
StatePublished - Jan 17 1995

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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