Re-engineering cytochrome P450 2B11dH for enhanced metabolism of several substrates including the anti-cancer prodrugs cyclophosphamide and ifosfamide

Ling Sun, Chong S. Chen, David J. Waxman, Hong Liu, James R. Halpert, Santosh Kumar

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

Based on recent directed evolution of P450 2B1, six P450 2B11 mutants at three positions were created in an N-terminal modified construct termed P450 2B11dH and characterized for enzyme catalysis using five substrates. Mutant I209A demonstrated a 3.2-fold enhanced kcat/Km for 7-ethoxy-4-trifluoromethylcourmarin O-deethylation, largely due to a dramatic decrease in Km (0.72 μM vs. 18 μM). I209A also demonstrated enhanced selectivity for testosterone 16β-hydroxylation over 16α-hydroxylation. In contrast, V183L showed a 4-fold increased kcat for 7-benzyloxyresorufin debenzylation and a 4.7-fold increased kcat/Km for testosterone 16α-hydroxylation. V183L also displayed a 1.7-fold higher kcat/Km than P450 2B11dH with the anti-cancer prodrugs cyclophosphamide and ifosfamide, resulting from a ∼4-fold decrease in Km. Introduction of the V183L mutation into full-length P450 2B11 did not enhance the kcat/Km. Overall, the re-engineered P450 2B11dH enzymes exhibited enhanced catalytic efficiency with several substrates including the anti-cancer prodrugs.

Original languageEnglish (US)
Pages (from-to)167-174
Number of pages8
JournalArchives of Biochemistry and Biophysics
Volume458
Issue number2
DOIs
StatePublished - Feb 15 2007

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Ifosfamide
Hydroxylation
Prodrugs
Metabolism
Cyclophosphamide
Cytochrome P-450 Enzyme System
Testosterone
Substrates
Neoplasms
Catalysis
Mutation
Enzymes

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology

Cite this

Re-engineering cytochrome P450 2B11dH for enhanced metabolism of several substrates including the anti-cancer prodrugs cyclophosphamide and ifosfamide. / Sun, Ling; Chen, Chong S.; Waxman, David J.; Liu, Hong; Halpert, James R.; Kumar, Santosh.

In: Archives of Biochemistry and Biophysics, Vol. 458, No. 2, 15.02.2007, p. 167-174.

Research output: Contribution to journalArticle

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