0000000000856031
AUTHOR
Claudia Pallotti
The use of trypsin to solubilize wall proteins from Candida albicans led to the identification of chitinase 2 as an enzyme covalently linked to the yeast wall structure
The use of trypsin to break proteins covalently linked to the yeast walls of Candida albicans released approx. 50% of the proteins, but also glucose and N-acetylglucosamine. Analysis by affinity chromatography indicated that glucose and/or N-acetylglucosamine formed part of the same supramolecular complexes with mannoproteins. These complexes would represent a new type of cell wall structuration in which beta-1,6 glucan and chitin are linked to proteins. An internal peptide from a 50-kDa protein released by trypsin was sequenced, showing 100% identity with chitinase 2 protein and 92% with chitinase 3. The electrophoretic mobility of the chitinase 2 protein was changed by treatment with Endo…
Cloning and characterization of the phenylalanyl-tRNA synthetase β subunit gene fromCandida albicans
A Candida albicans expression library was constructed from RNA isolated from regenerating protoplasts. A 1.4-kb cDNA clone was used to isolate a genomic fragment. Sequence analysis revealed an open reading frame of 593 amino acids with an overall identity of 63.6% with the phenylalanyl-tRNA synthetase beta subunit (FRS1) of Saccharomyces cerevisiae. We named it CaFRS1. It is located in a single copy in chromosome R, SfiI fragment M. Its expression showed a decrease during the cell wall regeneration process in protoplasts of both yeast and mycelial cells of C. albicans, suggesting its requirement thereof in initial steps of the cell wall synthesis.
Transglutaminase activity is involved in Saccharomyces cerevisiae wall construction
Transglutaminase activity, which forms the interpeptidic cross-link N(epsilon)-(gamma-glutamyl)-lysine, was demonstrated in cell-free extracts of Saccharomyces cerevisiae by incorporation of [(14)C]lysine into an exogenous acceptor, N,N'-dimethylcasein. Higher levels of the activity were present in the cell wall, which also contained endogenous acceptors. The enzyme activity in the wall was inhibited by cystamine, a known inhibitor of transglutaminase, and by EDTA, indicating a cation-dependent activity. After the endogenous wall acceptors were labelled radioactively by transglutaminase, extraction with SDS solubilized about 50% of the total radioactivity, while Zymolyase and chitinase each…