EFFECTS OF IONIC AND OSMOTIC STRENGTH ON THE GLUCOSYLTRANSFERASE OF RHIZOBIUM-MELILOTI RESPONSIBLE FOR CYCLIC BETA-(1,2)-GLUCAN BIOSYNTHESIS

The cyclic beta-(1,2)-glucans of Rhizobium meliloti and Agrobacterium tumefaciens play an important role during hypoosmotic adaptation, and the synthesis of these compounds is osmoregulated. Glucosyltransferase , the enzyme responsible for cyclic beta-(1,2)-glucan biosynthesis, is present constitutively, suggesting that osmotic regulation of the bio synthesis of these glucans occurs through modulation of enzyme activit y. In this study, we examined regulation of cyclic glucan biosynthesis in vitro with membrane preparations from R. meliloti. The results sho w that ionic solutes inhibit glucan synthesis, even when they are pres ent at low concentrations (e.g., 10 mM). In contrast, neutral solutes (glucose, sucrose, and the compatible solutes glycine betaine and treh alose) were found to stimulate glucan synthesis in vitro when they wer e present at high concentrations (e.g., 1 M). Furthermore, high concen trations of these neutral solutes were shown to compensate for the inh ibition of glucosyltransferase activity by ionic solutes. Consistent w ith their ionic character, the compatible solute potassium glutamate a nd the osmoprotectant choline chloride inhibited glucosyltransferase a ctivity in vitro. The results suggest that intracellular ion concentra tions, intracellular osmolarity and intracellular concentrations of no nionic compatible solutes all act as important determinants of glucosy ltransferase activity in vivo. Additional experiments were performed w ith an ndvA mutant defective for transport of cyclic glucans and an nd vB mutant that produces a C-terminal truncated glycosyltransferase. Cy clic beta-(1,2)-glucan biosynthesis, although reduced, was found to be osmoregulated in both mutants. These results reveal that NdvA and the C terminus of NdvB are not required for osmotic regulation of cyclic beta-(1,2)-glucan biosynthesis.