Substantial Spatial Flexibility and Hydrogen Bonding within the Catalysis Exerted by Cyclodextrin Artificial Glycosidases

Herein we report the synthesis of a novel 7(A),7(D)-dicyanohydrin-beta-cyclodextrin that catalyzes the hydrolysis of aryl glycosides with up to 5500 times rate increase (k(cat)/k(uncat)), functioning as a glycosidase enzyme mimic. For all glycoside substrates tested at 50 mm phosphate buffer this catalysis is superior to previously reported results for 6(A),6(D)-dicyanohydrin cyclodextrin (CD) artificial glycosidases, i.e. analogues which have their catalytic group one carbon atom closer to the cyclodextrin cavity. This provides proof of substantial flexibility within the catalysis exerted by these CD chemzymes. A series of permethylated mono- and dicyanohydrin alpha- and beta-CDs were also synthesized, and these showed more modest catalytic rate enhancements of up to 110 times (10% catalysis rate, relative to non-methylated analogues), implying that the permethylation blocks or hampers catalytically important binding between the saccharide part of the substrate and the CD. For comparison, the permethylated 6(A),6(D)-dicarboxylic acid beta-CD was also synthesized and afforded 25% activity (up to 250 times rate increase) relative to the nonpermethylated 6(A),6(D)-dicarboxylic acid beta-CD. This suggests that the catalytic effect of the polar interactions of the ionized carboxylate entity is less dependent on the substrate position. These findings afford new information on the scopes and boundaries for CD artificial glycosidase catalysis, and the spatial flexibility discovered fosters optimism for future advances and discoveries within the field of artificial enzymes.