SUBSTRATE RECOGNITION BY AMYLOGLUCOSIDASE - EVALUATION OF CONFORMATIONALLY BIASED ISOMALTOSIDES

Amyloglucosidase catalyzes the hydrolysis of methyl beta-maltoside (1) 30-50 times more rapidly than methyl alpha-isomaltoside (2). It is es tablished that OH-6', OH-4' and OH-3 are intimately involved in the hy drolysis of the maltoside whereas it is OH-6', OH-4', and OH-4 which a re involved in key polar interactions with the enzyme in the case of i somaltoside. Conformational analyses based on HSEA calculations indica te that the dispositions in space of OH-3 of maltose relative to OH-4' and OH-6' in the preferred conformation for the maltoside (1) is ener getically more readily achieved by methyl 6R-C-methyl-alpha-isomaltosi de (3), than for its 6-S-isomer (4). A kinetic evaluation of the hydro lysis in fact has shown that the S-compound is more strongly bound by the enzyme (K-m = 0.9 mM) than the parent isomaltoside (K-m = 24.5 mM) , whereas the S-compound has the weakest enzyme binding (K-m = 90 mM). Since the k(cat) values were all within the range 0.85 +/- 0.20 s(-1) , it is evident that the relative rates of hydrolysis are related to t he relative ease for the compounds to achieve an interaction of a hydr oxyl group in the aglycon of an alpha-D-glucopyranoside with the enzym e for the formation of the enzyme-substrate complex. The relative rate s of hydrolysis of the alpha-glucosides of the 1,3-dihydroxy-trans-dec alins, 5 and 6, provide further support for this highly desirable but not necessary recognition for the orientation of the reducing glucose unit in the active site.