Journal article
Active Site Mutations of Cytochrome P450cam Alter the Binding, Coupling, and Oxidation of the Foreign Substrates (R)- and (S)-2-Ethylhexanol
Archives of biochemistry and biophysics, Vol.398(2), pp.188-197
02/15/2002
Handle:
https://hdl.handle.net/2376/115503
PMID: 11831849
Abstract
Three factors are of primary importance with respect to designing efficient P450 biocatalysts. (1) The substrate must be oxidized at a significant rate. (2) The regioselectivity must heavily favor the desired product. (3) The enzyme must use the majority of the reducing equivalents from NADH or NADPH to produce product. The reaction we chose to study was oxidation of 2-ethylhexanol to 2-ethylhexanoic acid by P450cam. We examined four active site mutations: F87W, Y96W, T185F, and L244A. The mutations were chosen to improve 2-ethyhexanoic acid production by decreasing active site volume, increasing active site hydrophobicity, and improving stereoselectivity. The F87W and Y96W mutations improved regioselectivity, giving almost exclusively the desired product. The T185F mutation improved coupling of NADH to product formation. The L244A mutation altered the stereoselectivity of 2-ethylhexanoic acid production. These results indicate that active site mutations of P450cam can alter catalysis of 2-ethylhexanol.
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Details
- Title
- Active Site Mutations of Cytochrome P450cam Alter the Binding, Coupling, and Oxidation of the Foreign Substrates (R)- and (S)-2-Ethylhexanol
- Creators
- Kevin J French - Toxicology Training Program, Department of Environmental Medicine, University of Rochester School of Medicine and Dentistry, Rochester, New York, 14642Dan A Rock - Department of Chemistry, Washington State University, Pullman, Washington, 99164Denise A Rock - Department of Chemistry, Washington State University, Pullman, Washington, 99164John I Manchester - ArQule, Inc. Woburn, Massachusetts, 01801Barry M Goldstein - Department of Environmental Medicine, Department of Biochemistry and Biophysics, University of Rochester School of Medicine and Dentistry, Rochester, New York, 14642Jeffrey P Jones - Department of Chemistry, Washington State University, Pullman, Washington, 99164
- Publication Details
- Archives of biochemistry and biophysics, Vol.398(2), pp.188-197
- Academic Unit
- Chemistry, Department of
- Publisher
- Elsevier Inc
- Identifiers
- 99900547531901842
- Language
- English
- Resource Type
- Journal article