(1-->3)-beta-d-Glucan Synthase from Sugar Beet : I. Isolation and Solubilization

Abstract

A (1-->3)-beta-glucan synthase has been isolated from petiole tissue of sugar beet (Beta vulgaris L.). Enzyme activity is associated with a membrane fraction with a density of 1.03 grams per cubic centimeter when subjected to isopycnic density gradient centrifugation in Percoll. The reaction product was determined to be a linear (1-->3)-beta-glucan by methylation analysis and by glucanase digestion. (1-->3)-beta-Glucan synthase activity is markedly stimulated by Ca(2+); activation is half-maximal at about 50 micromolar Ca(2+) and is nearly saturated at 100 micromolar. Other divalent cations tested, Mg(2+), Mn(2+), and Sr(2+), also stimulate enzyme activity but are less effective. Enzyme activity was also stimulated up to 12-fold by beta-glucosides. Sirofluor, the fluorochrome from aniline blue, inhibited enzyme activity 95% when included at 1 millimolar. The enzyme was solubilized in Zwittergent 3-14; 85% of total enzyme activity was solubilized in 0.03% detergent and the optimal detergent-to-protein ratio was 0.3 at 3 milligrams per milliliter protein.