title: Challenges in assignment of allosteric effects in cytochrome P450-catalyzed substrate oxidations to structural dynamics in the hemoprotein architecture.
abstract: Cytochrome P450s (CYP) represent a superfamily of b-type hemoproteins catalyzing NAD(P)H-dependent oxidative biotransformation of a vast array of natural and xenobiotic compounds. Many eu- and prokaryotic members of this class of monooxygenases display complex non-Michaelis-Menten saturation kinetics, suggestive of homo-/heterotropic cooperativity arising from substrate-/effector-induced allosteric interactions. Here, the paradigm of multiple-ligand occupancy of the catalytic pocket in combination with enzyme oligomerization provides the most favored explanations for the atypical kinetic patterns. Making use of available data from crystallographic analyses, homology modeling and site-directed mutagenesis, the present review focuses on assessment of the topology of prospective key players dictating allosterism. Based on a general, CYP3A4-related construct, the majority of determinants were found to cluster within the six known substrate recognition sites (SRSs). Here, the B'/B'-C domains (SRS-1) and the F-helical region (SRS-2) harbor 51% of the critical residues, while SRS-4/5/6 each accommodate about 11-17% of the presumed docking spots. Of note, 12% of the total number of functional amino acids resides in non-SRS motifs. Average frequency of conservation of the allosteric sites examined was found to be fairly low (~13%), hinting at the requirement of some degree of conformational flexibility. Reactivity toward ligands coincides with the lipophilicity/hydrophilicity profile and bulkiness of the elements acting as selective filters. In sum, cooperative scenarios mainly pertain to regulative effects on substrate ingress, tuning of the open/closed equilibrium of the substrate access channel, modulation of the active-site capacity and productive ligand orientation toward the iron-oxene core. Deeper insight into the molecular mechanism of allostery may help avoid undesired drug-drug interplay in medicinal therapy and offer detoxification response to toxic agents. Finally, genetic manipulation of the cooperative machinery of P450s may provide a powerful tool in drug development.
authors.affiliation: Walther-Straub-Institut fuer Pharmakologie und Toxikologie, Goethestrasse 33, D-80336 Muenchen, Germany. Electronic address: hlavica@lrz.uni-muenchen.de.
authors.forename: Peter
authors.initials: P
authors.lastname: Hlavica
mesh_terms: D000494:Allosteric Regulation; D000818:Animals; D002384:Catalysis; D051544:Cytochrome P-450 CYP3A / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism*; D015195:Drug Design; D006801:Humans; D007501:Iron / Q000737:chemistry / Q000378:metabolism; D056004:Molecular Dynamics Simulation; D010084:Oxidation-Reduction; D013329:Structure-Activity Relationship
chemical_list: D007501:Iron; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human
keywords: Allosteric effects; Cooperativity; Cytochrome P450; Key determinants; Molecular modeling