Experiments with synchronized cultures have confirmed previous indications that the yeast chitin synthetases are regulated at the post- translational level, although regulation by synthesis and inactivation may also play a role in the case of chitin synthetase 2. Progressive deletions in the CHS2 gene have defined a sequence that is critical for activity and function of the corresponding chitin synthetase. A hybrid gene containing that sequence linked to a portion of the CHS1 gene led to expression of a Chs1-like synthetase that has lost the zymogenic character. beta(1->)glucan is the major structural component of the yeast cell wall. Purification of a component of beta(1->)glucan synthetase followed by photolabeling led to the identification of a 20 Kda protein as the GTP-binding subunit of this fraction. Previous work has suggested that the components of the yeast cell wall are covalently linked to each other, in particular that chitin is attached to beta(1->3)glucan. Digestion of yeast cell walls with beta glucanase and chitinase followed by reduction with borotritide and chromatography on size columns led to the isolation of oligosaccharides containing both N-acetylglucosamine and glucose residues. These oligosaccharides contain the previously postulated linkage between chitin and glucan. Because the oligosaccharides are absent in a chitin synthetase 3-deficient strain, it is concluded that chitin synthetase 3 catalyzes the synthesis of the chitin that is attached to glucan.
