Tryptophan‐96 in cytochrome P450 BM3 plays a key role in enzyme survival. Issue 1 (27th October 2022)

• Record Type: Journal Article
• Authors: Ravanfar, Raheleh; Sheng, Yuling; Gray, Harry B.; Winkler, Jay R.
• Is Part Of: FEBS letters. Volume 597:Issue 1(2023)
• Page Start: 59
• Page End: 64
• Publication Date: 2022-10-27
• Subjects: cytochrome P450 -- electron transfer -- total turnover number -- tryptophan -- tyrosine; Biochemistry -- Periodicals; Biophysics -- Periodicals; Molecular biology -- Periodicals; Biochimie -- Périodiques; Biochemistry; Biophysics; Molecular biology; Periodicals; 572.05
• DOI: 10.1002/1873-3468.14514
• Languages: English
• ISSNs: 0014-5793

Abstract

Abstract : Flavocytochrome P450 from Bacillus megaterium (P450BM3 ) is a natural fusion protein containing reductase and heme domains. In the presence of NADPH and dioxygen the enzyme catalyses the hydroxylation of long‐chain fatty acids. Analysis of the P450BM3 structure reveals chains of closely spaced tryptophan and tyrosine residues that might serve as pathways for high‐potential oxidizing equivalents to escape from the heme active site when substrate oxidation is not possible. Our investigations of the total number of enzyme turnovers before deactivation have revealed that replacement of selected tryptophan and tyrosine residues with redox inactive groups leads to a twofold reduction in enzyme survival time. Tryptophan‐96 is critical for prolonging enzyme activity, suggesting a key protective role for this residue. Abstract : Cytochrome P450BM3 hydroxylates an average of 340 p ‐nitrophenoxydodecanoic acid molecules before the enzyme is deactivated. Removal of selected tryptophan and tyrosine residues has little impact on enzyme activity but reduces total turnover by as much as one half. The results suggest that tryptophan and tyrosine protect cytochrome P450BM3 from damage during uncoupled turnover.