Chitin and Yeast Budding

The product was characterized as chitin by its insolubility in alkali, the release of glucosamine on acid hydrolysis, and the liberation of diacetylchitobiose and acetylglucosamine following stepwise enzymatic hydrolytic, concluding that chit in synthetase is not located outside the cytoplasmic membrane.

Abstract

Yeast cells were extracted with alkali and acetic acid according to the method of Bacon et al. (Biochem. J., 101, 36C (1966)). Examination of the extracted cell ghosts under the electron microscope revealed very thin cell envelopes with prominent bud scars in the shape of a shallow crater with a raised rim. The extracted cell ghosts, which had a high chitin content (15%), were treated with purified chitinase and glucanase. Electron micrographs showed that chitinase treatment did not destroy the integrity of the cell envelope, but largely eliminated the bud scar rims. On the other hand, after incubation with glucanase only fragments in the shape of bud scars, isolated or in groups, were observed. It is concluded that chitin is localized in a ring around the bud scar, sandwiched between two layers of glucan. The possible function of chitin during budding is discussed on the basis of this localization. During extraction of the yeast cells, a glucan fraction was isolated, which is soluble above pH 10, but rapidly forms a gel upon neutralization. This glucan was used to adsorb and purify the glucanase.