title: Conformational landscape and the selectivity of cytochrome P450cam.
abstract: Conformational heterogeneity and dynamics likely contribute to the remarkable activity of enzymes but are challenging to characterize experimentally. These features are of particular interest within the cytochrome P450 class of monooxygenases, which are of great academic, medicinal, and biotechnological interest as they recognize a broad range of substrates, such as various lipids, steroid precursors, and xenobiotics, including therapeutics. Here, we use linear and 2D IR spectroscopy to characterize the prototypical P450, cytochrome P450cam, bound to three different substrates, camphor, norcamphor, or thiocamphor, which are hydroxylated with high, low, and intermediate regioselectivity, respectively. The data suggest that specific interactions with the substrate drive the population of two different conformations, one that is associated with high regioselectivity and another associated with lower regioselectivity. Although Y96 mediates a hydrogen bond thought necessary to orient the substrate for high regioselectivity, the population and dynamics of the conformational states are largely unaltered by the Y96F mutation. This study suggests that knowledge of the conformational landscape is central to understanding P450 activity, which has important practical ramifications for the design of therapeutics with optimized pharmacokinetics, and the manipulation of P450s, and possibly other enzymes, for biotechnological applications.
authors.affiliation: Department of Chemistry, Indiana University, Bloomington, Indiana 47405, United States.
authors.forename: Edward J
authors.initials: EJ
authors.lastname: Basom
authors.affiliation: Department of Chemistry, Indiana University, Bloomington, Indiana 47405, United States.
authors.forename: James W
authors.initials: JW
authors.lastname: Spearman
authors.affiliation: Department of Chemistry, Indiana University, Bloomington, Indiana 47405, United States.
authors.forename: Megan C
authors.initials: MC
authors.lastname: Thielges
mesh_terms: D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry* / Q000378:metabolism*; D006860:Hydrogen Bonding; D008958:Models, Molecular; D009636:Norbornanes / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D055672:Static Electricity; D013379:Substrate Specificity
chemical_list: D009636:Norbornanes; D003577:Cytochrome P-450 Enzyme System