Abstract
A (1→3),(1→4)-β-glucan synthase (GTase) catalyzing the synthesis of (1→3),(1→4)-β-glucan was investigated using microsomal membranes prepared from developing barley (Hordeum vulgare L. cv. Shikokuhadaka 97) endosperms harvested 21 days after flowering. The microsomal fraction transferred [14C]Glc from UDP-[14C]Glc into endogenous acceptors, leading to the formation of (1→3),(1→4)-β-glucan. The production of (1→3),(1→4)-β-glucan was ascertained by specific enzymatic digestion with endo-(1→3),(1→4)-β-glucanase (lichenase), which released a radiolabeled trisaccharide (3-O-β-cellobiosyl-glucose) and a tetrasaccharide (3-O-β-cellotriosyl-glucose). The activity of the GTase was maximal at pH 9.0 and at 25°C and in the presence of at least 2mM Mg2+ (specific activity, 300 pmol/min/mg protein). Investigating the dependence of GTase activity on developmental stage (7 to 35 days after flowering) of the endosperms, we found an increase of activity during the initial development reaching a maximum at 19 days, followed by a gradual decrease as the endosperms matured.
