How do perfluorinated alkanoic acids elicit cytochrome P450 to catalyze methane hydroxylation? An MD and QM/MM study

Recent experimental studies show that the usage of perfluoro decanoic acid (PFDA), as a dummy substrate, can elicit P450BM₃ to perform hydroxylation of small alkanes, such as methane (Angew. Chem., Int. Ed., 2011, 50, 2720) and propane (Angew. Chem., Int. Ed., 2011, 50, 2720 and Angew. Chem., Int. Ed., 2011, 50, 5315). To comprehend the mechanism whereby PFDA operates to potentiate P450BM₃ to catalyze the hydroxylation of small alkanes, we used molecular dynamics (MD) and hybrid quantum mechanical/molecular mechanical (QM/MM) calculations. The MD results show that without the PFDA, methane escapes the active site, while the presence of PFDA can potentially induce a productive Cpd I-Methane juxtaposition for rapid oxidation. Nevertheless, when only a single methane molecule is present near the PFDA, it still escapes the pocket within less than a nanosecond. However, when three methane molecules are present in the pocket, they alternate quasi-periodically such that at all times (within 10 ns), a molecule of methane is always present in the proximity of Cpd I in a reactive conformation. Our results further demonstrate that the PFDA does not exert any electrostatic catalysis, whether the PFDA is in the protonated or deprotonated forms. Taken together, we conclude that methane hydroxylation requires, in addition to PFDA, a high partial pressure of methane that will cause a high methane concentration in the active site. Further study of ethane and propane hydroxylations demonstrates that higher alkane concentration is helpful for all the three small alkanes. Thus for the smallest alkane, methane, at least three molecules are necessary, whereas for the larger ethane, two molecules are needed to force one ethane to be closer to Cpd I. Finally, for propane, a second molecule is helpful but not absolutely necessary; for this molecule the PFDA may well be sufficient to keep propane close to Cpd I for efficient oxidation. We therefore propose that high alkane pressure should assist small alkane hydroxylation by P450 in a manner inversely proportional to the size of the alkanes.