Tracking Spin Densities In P450 Mutants

Cytochrome P450 is a family of powerful enzymes commonly found in the human body responsible for substrate (drugs) hydroxylation. An essential player in detoxification, should the protein's mechanism be elucidated, more biocompatible drugs could be produced. Currently, two theories exist on the locus of P450 spin density; one argues a concentration at the heme iron and porphyrin group, while the other argues a dispersion across heme propionates and proximal residues. Given the vast utility which may result from greater understanding, a computational study of P450 and its mutants was embarked upon by use of mixed quantum mechanics and molecular mechanics (QM/MM) methods.

First, a crystal structure of P450 from pseudomonas putida complexed with camphor was loaded from the Protein Data Bank. Two mutants of said protein were generated by point mutation at residue 297 (in the wild type as aspartic acid, “ASP”) to a sparagine (“ASN ” ) and to leucine (“ LEU”). Next, conformation changes were determined by a rotamer library protein structure prediction program, PLOP. Then QM/MM run found atomic charges, and molecular dynanics used those charges to simulate protein activity at body temperature. (The combined results determined charge and spin densities). Lastly, a repeat QM/MM run on selected frames generated by molecular dynanics discovered any changes in charge and spin densities. With regard to ASN and LEU mutants, the process was essentially the same. In addition, following PLOP loop sampling, further loop sampling was performed on residues in proximity to residue 297 to check for conformation changes.