Direct Experimental Evidence for the High Chemical Reactivity of α‐ and β‐Xylopyranosides Adopting a ^2,5B Conformation in Glycosyl Transfer

Abstract

The effect of a (2,5)B boat conformation on xyloside reactivity has been investigated by studying the hydrolysis and glycosylation of a series of synthetic xyloside analogues based on a 2-oxabicyclo[2.2.2]octane framework, which forces the xylose analogue to adopt a (2,5)B conformation. The locked β-xylosides were found to hydrolyze 100-1200 times faster than methyl β-D-xylopyranoside, whereas the locked α-xylosides hydrolyzed up to 2×10(4) times faster than methyl α-D-xylopyranoside. A significant rate enhancement was also observed for the glycosylation reaction. The high reactivity of these conformers can be related to the imposition of a (2,5)B conformation, which approximates a transition state (TS) boat conformation. In this way, the energy penalty required to go from the chair to the TS conformation is already paid. These results parallel and support the observation that the GH-11 xylanase family force their substrate to adopt a (2,5)B conformation to achieve highly efficient enzymatic glycosidic bond hydrolysis.

Exploration of conformational flexibility and hydrogen bonding of xylosides in different solvents, as a model system for enzyme active site interactions

“…It was further established that the suggested conformers made the molecule at hand more apt to undergo reaction, e.g., increasing the tendency for aglycon dissociation by increasing the C1-O1 bond distance in mannose upon a transition from the 4 C 1 to the 1 S 5 ring conformation. Recent experiments performed with xylose analogues locked in the 2,5 B conformation 4 displayed enhanced glycolysis and glycosylation rates, thus confirming these calculations.…”

How Sugars Pucker: Electronic Structure Calculations Map the Kinetic Landscape of Five Biologically Paramount Monosaccharides and Their Implications for Enzymatic Catalysis

“…Amorim et al tested the hypothesis that the 2,5 B conformation is especially amenable to catalysis. 91 They compared hydrolysis rates of a xyloside and a bicyclic xyloside analogue locked into a 2,5 B geometry by a C−C bridge between C2 and C5. They found hydrolysis rates for the locked 2,5 B xyloside to be at least 2 orders of magnitude faster than the flexible xyloside.…”

“…It may be that the higher C1 partial charge offers more catalytic benefit, making it worth overcoming the higher free energy. Amorim et al tested the hypothesis that the 2,5 B conformation is especially amenable to catalysis . They compared hydrolysis rates of a xyloside and a bicyclic xyloside analogue locked into a 2,5 B geometry by a C–C bridge between C2 and C5.…”

The complete conformational free energy landscape of β-xylose reveals a two-fold catalytic itinerary for β-xylanases

“… 24 Support for the second itinerary ( 2 S O → [ 2,5 B] ǂ → 5 S 1 ) mainly originates from reports on covalently bound substrates of family 11 GHs in which the xylose saccharide located at the –1 subsite (2-deoxy-2-fluoroxylopyranosyl substrate) adopts a 2,5 B conformation, 25 , 26 as well as experiments on synthetic xyloside analogues locked on a 2,5 B conformation. 27 Computational modeling studies on B. circulans GH11 xylanase addressed this question, concluding that the conformation of the xylose saccharide in the Michaelis complex is either 2 S O or 2,5 B. 28 , 29 To add to the confusion, a recent structural study reports an almost undistorted conformation ( 4 C 1 / O E) for a xylohexaose substrate in complex with a GH11 xylanase mutant, predicting an unusual O E → [ O S 2 ] ǂ → B 2,5 itinerary ( Fig.…”