Biosynthesis of glucans in mung bean seedlings: Formation of β-(1 → 4)-glucans from GDP-glucose and β-(1 → 3)-glucans from UDP-glucose

Abstract

A particulate enzyme system from Phaseolus aureus catalyzed the incorporation of radioactivity from UDP-glucose- 14C and GDP-glucose- 14C into alkali-soluble and alkali-insoluble polysaccharides. Most (90%) of the radioactivity incorporated from UDP-glucose- 14C was in the water-insoluble, alkali-soluble polymers while the remaining 10% was in the alkali-insoluble fraction. On the other hand, most (60–90%) of the 14C incorporated from GDP-glucose- 14C remained in the alkali-insoluble polysaccharides. The incorporation of both UDP-glucose- 14C and GDP-glucose- 14C into the alkali-soluble and alkali-insoluble materials reached a maximum in 1 to 2 min and then leveled off, but incorporation of both nucleotides was linear with protein concentration. The radioactive, alkali-insoluble polysaccharides synthesized from both UDP-glucose- 14C and GDP-glucose- 14C were compared by various criteria to determine the nature of the polymers and of their glycosidic linkages. Acetolysis or partial acid hydrolysis of the GDP-glucose- 14C synthesized product gave rise to cellobiose as the only 14C-disaccharide whereas the product formed from UDP-glucose- 14C produced mostly laminaribiose (β, 1 → 3). Periodate oxidation of either the larger oligosaccharides or the insoluble polymer formed from GDP-glucose- 14C, followed by reduction and hydrolysis, led to the formation of 14C-erythritol, confirming a 1 → 4 linkage. On the other hand, periodate oxidation of the UDP-glucose- 14C products under identical conditions produced mostly 14C-glucose, further suggesting a 1 → 3 linkage. The alkali-soluble polymers produced from UDP-glucose- 14C were identical to the alkali-insoluble polysaccharide on the basis of acetolysis and periodate oxidation. Treatment of the particles with digitonin resulted in their loss in the ability to incorporate radioactivity from UDP-glucose- 14C and GDP-glucose- 14C into polysaccharides. However, 70% of the activity with GDP-glucose- 14C, but virtually none of the UDP-glucose- 14C activity, was restored by adding 7 × 10 −3 m Mg 2+ to the particles. The UDP-glucose- 14C polysaccharide synthesizing activity was found in the digitonin-treated supernatant liquid. This soluble enzyme fraction catalyzed the incorporation of radioactivity from UDP-glucose- 14C into a water-insoluble polysaccharide which had the properties of aβ-(1 → 3)-glucan. Radioactive laminaribiose and smaller amounts of laminaritriose and laminaritetraose were also formed by the soluble enzyme.