Steric course of the hydration of D-gluco-octenitol catalyzed by .alpha.-glucosidases and by trehalase

Abstract

Crystalline Aspergillus niger .alpha.-glucosidase and highly purified preparations of rice .alpha.-glucosidase II and Trichoderma reesei trehalase were found to catalyze the hydration of [2-2H]-D-gluco-octenitol, i.e., (Z)-3,7-anhydro-1,2-dideoxy-[2-2H]-D-gluco-oct-2-enitol, to yield 1,2-dideoxy-[2-2H]-D-gluco-octulose. In each case, the stereochemistry of the reaction was elucidated by examining the newly formed centers of asymmetry at C-2 and C-3 of the hydration product. The C-1 to C-3 fragment of each isolated [2-2H]-D-gluco-octulose product was recovered as [2-2H]propionic acid and identified by its positive optical rotatory dispersion at the S isomer, showing that each enzyme had protonated the octenitol (at C-2) from above its reface. 1H NMR spectra of enzyme/D-gluco-octenitol digests in D2O showed that the .alpha.-anomer of [2-2H]-D-gluco-octulose was exclusively produced by each .alpha.-glucosidase, whereas the .beta.-anomer was formed by action of the trehalase. The trans hydration catalyzed by the .alpha.-glucosidases was found to be very strongly inhibited by the substrate; the cis hydration reaction catalyzed by the trehalase showed no such inhibition. Special importance is attached to the finding that in hydrating octenitol each enzyme creates a product of the same anomeric form as in hydrolyzing an .alpha.-D-glucosidic substrate. This result adds substantially to the growing evidence that individual glycosylases create the configuration of their reaction products by a means that is independent of donor substrate configuration, that is, by a means other than "retaining" or "inverting" substrate configuration.