title: 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.
abstract: 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 inactivates 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 stereoisomers 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 epoxide-based "suicide" inhibitors may be enhanced significantly by the use of inhibitors bearing only one stereoisomeric form of the epoxide group.
authors.affiliation: Department of Biochemistry, La Trobe University, Bundoora, Victoria, Australia.
authors.forename: P B
authors.initials: PB
authors.lastname: Høj
authors.forename: E B
authors.initials: EB
authors.lastname: Rodriguez
authors.forename: J R
authors.initials: JR
authors.lastname: Iser
authors.forename: R V
authors.initials: RV
authors.lastname: Stick
authors.forename: B A
authors.initials: BA
authors.lastname: Stone
mesh_terms: D001412:Bacillus subtilis / Q000201:enzymology; D001665:Binding Sites; D002475:Cellobiose / Q000031:analogs & derivatives* / Q000494:pharmacology; D004852:Epoxy Compounds / Q000737:chemistry*; D006026:Glycoside Hydrolases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D001467:Hordeum / Q000201:enzymology; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011487:Protein Conformation; D013237:Stereoisomerism
chemical_list: D004852:Epoxy Compounds; D007527:Isoenzymes; D002475:Cellobiose; D006026:Glycoside Hydrolases; C028020:licheninase