Substrate binding, productive-mode

The mechanism is in complete agreement with results from recent tryptophan fluorescence experiments (which, due to the inviolability of microscopic reversibility, also hold in the synthesis mode) that establish definitively that (i) P, cannot simply bind spontaneously, (ii) an enzyme species with all three catalytic sites occupied is the only catalytically competent species, and (iii) release of product and binding of substrate caimot be simultaneous, rather product release must precede substrate binding [38]. 

In some reactions, a substrate binds in an alternative unreactive mode at the active site of the enzyme, in competition with the productive mode of binding ... 

Nonproductive binding. The larger substrate binds in the productive mode only, but the smaller one, in addition to binding more weakly in the productive mode, binds in nonproductive modes, lowering the KM. The fccat is correspondingly lower. 

The substrate binding pocket of horse liver alcohol dehydrogenase comprises residues from both subunits (Fig. 26B) [123]. The active site is shown in Fig. 27, with NAD(H) bound, and p-bromobenzyl alcohol bound in a non-productive binding mode. The hydrophobic residues (from both subunits) that line the substrate binding... 

Enzymatic action can be defined on three levels operational kinetics, molecular architecture, and chemical mechanism. Operational kinetic data have given indirect information about cellulolytic enzyme mode of action along with important information useful for modeling cellulose hydrolysis by specific cellulolytic enzyme systems. These data are based on measurement of initial rates of enzyme hydrolysis with respect to purified celluloses and their water soluble derivatives over a range of concentrations of both substrate and products. The resulting kinetic patterns facilitate definition of the enzyme s mode of action, kinetic equations, and concentration based binding constants. Since these enable the enzymes action to be defined with little direct knowledge of its mechanistic basis, the rate equations obtained are referred to as operational kinetics. The rate patterns have enabled mechanisms to be inferred, and these have often coincided with more direct observations of the enzyme s action on a molecular level [2-4]. 

Hypoxanthine is oxidized at carbon 2 by both molybdenum hydroxylases, although xanthine oxidase is much more effective as a catalyst in this reaction [ 10]. A methyl substituent in this position prevents oxidation by either enzyme. Introduction of A-methyl substituents into the hypoxanthine nucleus produces dramatic effects on enzymic oxidation rates and also gives some insight into the productive modes of binding to each enzyme. Thus, it has been proposed that hypoxanthine tautomerizes in the xanthine oxidase-substrate complex to the 3-NH-form with a simultaneous shift of the NH-group in the imidazole ring from position 9 to 7 [ 198,200]. In support of this hypothesis, when tautomerism in the imidazole ring is prevented by substitution at N-7 or N-9, such compounds are almost refractory to oxidation (see Table 3.9)... 

---