Implication of histidine at the active site of exo-beta-(1-3)-D-glucanase from Basidiomycete sp. QM 806.
The enzyme, exo-beta-(1-3)-D-glucanase, (EC 3.2.1-) obtained from a culture filtrate of Basidiomycete sp. QM 806, has been obtained in a highly purified form and preliminary investigations on its mechanism of action have been reported (Peterson, D. R., and Kirkwood, S. (1975) Carbohydr. Res. 41, 273-283). Studies reported in this paper, have provided strong evidence for the role of histidine in the catalytic site of this carbohydrase. Chemical modifications of the amino acid residues in the enzyme with diazotized 5-amino-1H-tetrazole or tetranitromethane caused irreversible loss of enzyme activity which varied according to the time of exposure to, or concentration of the inhibitor. Prior incubation of the enzyme with a substrate considerably reduced the extent of this inhibition. Amino acid analysis of the enzyme treated in these ways clearly indicated that the substrate protected histidine residues from chemical modification by the diazotized 5-amino-1H-tetrazole. Chemical modification of both histidine and tyrosine residues were effected by incubating the enzyme with the inhibitors described above. Although evidence is presented to suggest that tyrosine is not directly involved in the active site of the enzyme (the catalytic site or the binding site), the role of this residue in the maintenance of the enzyme conformation is discussed. Enzyme assays carried out either in aqueous or deuterated buffer systems provided further evidence which is consistent with the proposed enzyme mechanism.
D000596:Amino Acids; D001487:Basidiomycota / Q000201:enzymology*; D001665:Binding Sites; D055598:Chemical Phenomena; D002621:Chemistry; D043326:Glucan 1,3-beta-Glucosidase; D005936:Glucans; D005959:Glucosidases / Q000378:metabolism*; D006639:Histidine; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D011134:Polysaccharides / Q000378:metabolism; D013329:Structure-Activity Relationship; D013777:Tetrazoles / Q000494:pharmacology; D001617:beta-Glucosidase / Q000037:antagonists & inhibitors / Q000378:metabolism*
D000596:Amino Acids; D005936:Glucans; D011134:Polysaccharides; D013777:Tetrazoles; D006639:Histidine; C008247:laminaran; D005959:Glucosidases; D001617:beta-Glucosidase; D043326:Glucan 1,3-beta-Glucosidase