Chain length-dependent cooperativity in fatty acid binding and oxidation by cytochrome P450BM3 (CYP102A1)
| Autor: | Stephen Bell, Benjamin C Rowlatt, Luet-Lok Wong, Anthony J F Strong, Christopher J. C. Whitehouse, Jake A. Yorke |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2016 |
| Předmět: |
Cytochrome
Palmitic Acid Cooperativity 010402 general chemistry Myristic Acid 01 natural sciences Biochemistry Palmitic acid Structure-Activity Relationship 03 medical and health sciences chemistry.chemical_compound Bacterial Proteins Cytochrome P-450 Enzyme System Fatty acid binding Drug Discovery Heme NADPH-Ferrihemoprotein Reductase 030304 developmental biology chemistry.chemical_classification 0303 health sciences biology Fatty Acids Osmolar Concentration Lauric Acids Fatty acid Cell Biology Phosphate Lauric acid 0104 chemical sciences chemistry biology.protein Biophysics Oxidation-Reduction Research Article Biotechnology |
| Popis: | Fatty acid binding and oxidation kinetics for wild type P450(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 50 mmol/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 (cat). The results suggest that phosphate has a specific effect in promoting the first electron transfer step, and that P450(BM3) could modulate Bacillus membrane morphology and fluidity via palmitate oxidation in response to the external phosphate concentration. |
| Databáze: | OpenAIRE |
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