title: The oxidative half-reaction of Old Yellow Enzyme. The role of tyrosine 196.
abstract: Tyrosine 196 in Old Yellow Enzyme (OYE) was mutated to phenylalanine, and the resulting mutant enzyme was characterized to evaluate the mechanistic role of the residue. The residue demonstrates little effect on ligand binding and the reductive half-reaction, but a dramatic slowing by nearly 6 orders of magnitude of its oxidative half-reaction with 2-cyclohexenone. Observation of the oxidative half-reaction with a series of substrates allows us to propose a model describing the mechanism of the oxidative half-reaction. In addition, the curtailed reactivity with enones allows for characterization of the manner in which reduced enzyme primes the substrate for the redox reaction by observation of the Michaelis complex with reduced enzyme bound to substrate.
authors.affiliation: Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor, Michigan 48109-0606, USA.
authors.forename: R M
authors.initials: RM
authors.lastname: Kohli
authors.forename: V
authors.initials: V
authors.lastname: Massey
mesh_terms: D000171:Acrolein / Q000031:analogs & derivatives / Q000737:chemistry; D001665:Binding Sites; D002384:Catalysis; D003512:Cyclohexanones / Q000737:chemistry; D007700:Kinetics; D016296:Mutagenesis; D009252:NADPH Dehydrogenase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010084:Oxidation-Reduction; D011487:Protein Conformation; D014443:Tyrosine / Q000378:metabolism*
chemical_list: D003512:Cyclohexanones; D014443:Tyrosine; C028215:2-cyclohexen-1-one; D000171:Acrolein; D009252:NADPH Dehydrogenase; C012843:cinnamaldehyde