Kinetics of befunolol reductase from rabbit liver

The kinetic mechanism for the reduction of befunolol catalyzed by befunolol reductase from rabbit liver was investigated. From the initial velocity analysis, product inhibition and coenzyme binding studies, the reduction of befunolol was found to proceed through an ordered Bi Bi mechanism, in which beta-nicotinamide adenine dinucleotide phosphate, reduced form (NADPH) binds to the enzyme firstly and NADP+ leaves lastly. NADPH bound to the free enzyme at a molar ratio of 1:1. Furthermore, the result of dead-end inhibition by Cibacron blue F3GA, a nucleotide analogue which binds to many enzymes, was consistent with the ordered Bi Bi mechanism for the enzyme.