Roles of key active-site residues in flavocytochrome P450 BM3.

@article{Noble1999RolesOK,
  title={Roles of key active-site residues in flavocytochrome P450 BM3.},
  author={Michael A. Noble and Caroline S. Miles and Stephen K. Chapman and Dominikus A Lysek and Angela C. Mackay and Graeme A. Reid and Robert P. Hanzlik and Andrew W. Munro},
  journal={The Biochemical journal},
  year={1999},
  volume={339 ( Pt 2)},
  pages={
          371-9
        }
}
The effects of mutation of key active-site residues (Arg-47, Tyr-51, Phe-42 and Phe-87) in Bacillus megaterium flavocytochrome P450 BM3 were investigated. Kinetic studies on the oxidation of laurate and arachidonate showed that the side chain of Arg-47 contributes more significantly to stabilization of the fatty acid carboxylate than does that of Tyr-51 (kinetic parameters for oxidation of laurate: R47A mutant, Km 859 microM, kcat 3960 min-1; Y51F mutant, Km 432 microM, kcat 6140 min-1; wild… Expand
Structural determinants of active site binding affinity and metabolism by cytochrome P450 BM-3.
TLDR
Results indicate that binding affinity and catalytic turnover are fatty acid carbon-chain length dependent, and that the catalytic efficiency and the regioselectivity of fatty acid metabolism by BM-3 are determined by active site binding coordinates that control acceptor carbon orientation and proximity to the heme iron. Expand
A single active-site mutation of P450BM-3 dramatically enhances substrate binding and rate of product formation.
TLDR
Close examination of the crystal structures of the substrate-bound native and A328V mutant BM-3 BMPs indicates that the positioning of the substrates is essentially identical in the two forms of the enzyme, with the two valine methyl groups occupying voids present in the active site of the wild-type substrate- bound structure. Expand
The role of Thr268 and Phe393 in cytochrome P450 BM3.
TLDR
An important role is suggested for this active site threonine in substrate recognition and in maintaining an efficiently functioning enzyme. Expand
Probing the active site of flavocytochrome P450 BM3
The self sufficient class HI monooxygenase, flavocytochrome P450 BM3, from Bacillus megaterium, is a heme containing redox enzyme that catalyses the hydroxylation of long-chain fatty acids.Expand
Filling a hole in cytochrome P450 BM3 improves substrate binding and catalytic efficiency.
TLDR
The crystal structure of the haem domain of the Ala82Phe and Ala82Trp mutants are determined, indicating that the tighter binding in the mutant reflects a shift in the conformational equilibrium of the substrate-free enzyme towards the conformation seen in the substrate complex rather than differences in the enzyme-substrate interactions. Expand
Role of Leu188 in the Fatty Acid Hydroxylase Activity of CYP102A1 from Bacillus megaterium
Abstract P450 BM3 (CYP102A1) from Bacillus megaterium catalyzes the subterminal hydroxylation of fatty acids with 12-22 carbons at the ω-1, ω-2 and ω-3 positions. Several amino acids located at theExpand
ROLE OF RESIDUE 87 IN SUBSTRATE- AND REGIOSELECTIVITY OF DRUG METABOLIZING CYTOCHROME P450 BM3 M11
BM3, originating from Bacillus megaterium, is a highly active enzyme which has attracted much attention because of its potential applicability as a biocatalyst for oxidative reactions. Previously weExpand
Flavocytochrome P450 BM3 Mutant A264E Undergoes Substrate-dependent Formation of a Novel Heme Iron Ligand Set*
TLDR
A novel charge-transfer transition in the near-infrared magnetic CD spectrum provides a spectroscopic signature characteristic of the new A264E hemeIron ligation state, indicating that structural rearrangements occur following heme iron reduction to allow dioxygen binding. Expand
Effect of Mutation and Substrate Binding on the Stability of Cytochrome P450BM3 Variants.
TLDR
It is demonstrated that the heme domain R47L/F87V/L188Q/E267V/F81I pentuple mutant (PM) is destabilized because of the disruption of hydrophobic contacts and salt bridge interactions, and the lid domain, β-sheet 1, and Cys ligand loop are the most labile regions and, thus, potential sites for stabilizing mutations. Expand
Functional characterisation of an engineered multidomain human P450 2E1 by molecular Lego
TLDR
Results of a fusion between a human P450 enzyme and a bacterial reductase that for the first time is shown does not require the addition of lipids or detergents to achieve wild-type-like activities are presented. Expand
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References

SHOWING 1-5 OF 5 REFERENCES
Cytochromes P450: structure, function and mechanism.
Part 1 Introduction: historical background isolation, purification and characterization spectral and related physiochemical properties substrate-binding spectra polarized optical spectroscopyExpand
Cytochrome P-450: Structure, Mechanism, and Biochemistry
TLDR
The Cytochrome P450 and the Metabolism and Bioactivation of Arachidonic Acid and Eicosanoids in Plants and the Diversity and Importance of Microbial Cytochromes P450 are studied. Expand
Cytochrome P450
TLDR
It is shown that in allylic hydroxyl ations (catalyzed by either model metalloporphyrins or cytochrome P450) rehydridization could occur to yield multiple products. Expand
ed.), pp. 575–606
  • Narhi, L.-O. and Fulco, A. J
  • 1987