Active site-directed inhibition by optically pure epoxyalkyl cellobiosides reveals differences in active site geometry of two 1,3-1,4-beta-D-glucan 4-glucanohydrolases. The importance of epoxide stereochemistry for enzyme inactivation
Høj, P.B.; Rodriguez, E.B.; Iser, J.R.; Stick, R.V.; Stone, B.A.
Journal of Biological Chemistry 266(18): 11628-11631
1991
ISSN/ISBN: 0021-9258
PMID: 1904865
Accession: 006983408
1,3-1,4-.beta.-D-Glucan 4-glucanohydrolases (EC 3.2.1.73) from Bacillus subtilis and barley (Hordeum vulgare) with identical substrate specificities but unrelated primary structures have been probed with (R,S)-epoxyalkyl (-propyl, -butyl, -pentyl) .beta.-cellobiosides and with optically pure (3S)- and (3R)-3,4-cellobiosides as active site-directed inhibitors. The optimal aglycon length for inactivation differs for the two enzymes, and they are differentially inhibited by the pure epoxybutyl .beta.-cellobioside diastereoisomers. The (3S)-epoxybutyl .beta.-cellobioside inactivities the B. subtilis enzyme much more efficiently than does the (3R)-isomer, whereas the reverse is true for the barley enzyme. Both enzymes are inactivated by a mixture of the steroeoisomerase at a rate intermediate of that observed with the individual isomers. The two .beta.-glucan endohydrolases may therefore employ sterically different mechanisms to achieve glycoside bond hydrolysis in their common substrate. The efficiency and specificity of expoxide-based "sucicide" inhibitors may be enhanced significantly by the use of inhibitors bearing only one steroisomeric forn of the epoxide group.