On the Nature and Formation of the Fibrillar Nets Produced by Protoplasts of Saccharomyces cerevisiae in Liquid Media: An Electronmicroscopic, X-ray Diffraction and Chemical Study

Abstract

SUMMARYThe nets produced by protoplasts of Saccharomyces cerevisiae in liquid culture media consisted of microfibrils about 20 nm wide, forming flat, fairly straight bundles of variable width and length, up to about 500 nm wide and 4 pm long. Ends of microfibrils were seldom found. They were not attacked by chitinase or dilute acids, but the net structure disappeared in 3 % (w/v) NaOH, leaving about 60 % dry wt of the nets as-partly microfibrillar clusters. The X-ray powder pattern from the nets, in contrast to that from normal walls, exhibited a set of well-defined rings which identified two micro-crystalline constituents : chitin and unbranched chains of /3-(1+ +linked D-glucose residues. These latter were the alkali-soluble fraction. The X-ray diagram of the glucan, corresponding to that of paramylon, indicated an in vivo crystal modification. Up to 15 % dry wt was chitin which was formed de novo by the protoplasts.A fine net structure of microfibrils about 7.5 to 10 nm thick with meshes about 20 to 60 nm wide was demonstrated in normal walls, forming the entire inner layer and consisting mainly of yeast glucan. This glucan and chitin were only slightly crystalline in these walls. The features of the glucan and chitin of the protoplast nets indicate that enzymes active in normal wall formation were differentially removed or inactivated by the liquid medium.

Biosynthesis of Alkali-insoluble Cell-wall Glucan in Schizophyllum commune Protoplasts

“…A homopolymer of (1 -+ 3)-P-glucan is also formed on Saccharomyces cereuisiae protoplasts (Kreger & Kopecka, 1976) without the additional (1 + 6)-P-linked residues that are present in the normal walls of this yeast (Manners et al, 1973a, b ;Fleet & Manners, 1976). Similarly, experiments with isolated enzyme preparations from different fungi have yielded homopolymers of (1 --t 3)-P-glucans, whereas the alkali-insoluble P-glucans from the walls of the fungi contained a branched (1 -+ 3)-p/(l -, 6)-/?-glucan (Wang & Bartnicki Garcia, 1976;Shematek et al, 1980;Larriba et al, 198 1).…”

Architecture and Biosynthesis of the Saccharomyces cerevisiae Cell Wall

“…The order in which wall components are assembled has been inferred from analyses of the material formed when spheroplasts regenerate their walls and from the wall compositions of mutants unable to make a particular component (Kreger and Kopecká 1976;Roh et al 2002b). The starting component is b1,3-glucan, which is necessary for incorporation of both b1,6-glucan and mannoproteins.…”

Produção, purificação, clonagem e aplicação de enzimas líticas

“…Estudos da parede celular de leveduras com a utilização de microscópio eletrônico permitiram a Kreger e Kopecká 24 confirmarem a posição interna da fração de glucana.…”