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|Title:||Chain length-dependent cooperativity in fatty acid binding and oxidation by cytochrome P450BM3 (CYP102A1)|
|Citation:||Protein & Cell, 2011; 2(8):656-671|
|Publisher:||Gaodeng Jiaoyu Chubanshe|
|Benjamin Rowlatt, Jake A. Yorke, Anthony J. Strong, Christopher J. C. Whitehouse, Stephen G. Bell, Luet-Lok Wong|
|Abstract:||Fatty acid binding and oxidation kinetics for wild type P450(sub)BM3 (CYP102A1) from Bacillus megaterium have been found to display chain length-dependent homotropic behavior. Laurate and 13-methyl-myristate display Michaelis-Menten behavior while there are slight deviations with myristate at low ionic strengths. Palmitate shows Michaelis-Menten kinetics and hyperbolic binding behavior in 100 mmol/L phosphate, pH 7.4, but sigmoidal kinetics (with an apparent intercept) in low ionic strength buffers and at physiological phosphate concentrations. In low ionic strength buffers both the heme domain and the full-length enzyme show complex palmitate binding behavior that indicates a minimum of four fatty acid binding sites, with high cooperativity for the binding of the fourth palmitate molecule, and the full-length enzyme showing tighter palmitate binding than the heme domain. The first flavin-to-heme electron transfer is faster for laurate, myristate and palmitate in 100 mmol/L phosphate than in 50mmol/L Tris (pH 7.4), yet each substrate induces similar high-spin heme content. For palmitate in low phosphate buffer concentrations, the rate constant of the first electron transfer is much larger than k(sub)cat. The results suggest that phosphate has a specific effect in promoting the first electron transfer step, and that P450(sub)BM3 could modulate Bacillus membrane morphology and fluidity via palmitate oxidation in response to the external phosphate concentration.|
|Keywords:||P450BM3; monooxygenase; fatty acid; cooperativity; allosteric effect; CYP102A1|
|Rights:||© Higher Education Press and Springer-Verlag Berlin Heidelberg 2011|
|Appears in Collections:||Chemistry and Physics publications|
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