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Isolation of the catalase T structural gene of Saccharomyces cerevisiae by functional complementation.
- Source :
-
Molecular and cellular biology [Mol Cell Biol] 1983 Sep; Vol. 3 (9), pp. 1545-51. - Publication Year :
- 1983
-
Abstract
- The catalase T structural gene of Saccharomyces cerevisiae was cloned by functional complementation of a mutation causing specific lack of the enzyme (cttl). Catalase T-deficient mutants were obtained by UV mutagenesis of an S. cerevisiae strain bearing the cas1 mutation, which causes insensitivity of catalase T to glucose repression. Since the second catalase protein of S. cerevisiae, catalase A, is completely repressed on 10% glucose, catalase T-deficient mutant colonies could be detected under such conditions. A cttl mutant was transformed with an S. cerevisiae gene library in plasmid YEp13. Among the catalase T-positive clones, four contained overlapping DNA fragments according to restriction analysis. Hybridization selection of yeast mRNA binding specifically to one of the cloned DNAs, translation of this mRNA in cell-free protein synthesis systems, and demonstration of catalase T protein formation by specific immunoadsorption showed that the catalase T structural gene had been cloned. By subcloning, the gene was located within a 3.5-kilobase S. cerevisiae DNA fragment. As in wild-type cells, catalase T synthesis in cttl mutant cells transformed with plasmids containing this fragment is sensitive to glucose repression. By DNA-RNA hybridization, catalase T transcripts were shown to be present in oxygen-adapting cells but absent from heme-deficient cells.
Details
- Language :
- English
- ISSN :
- 0270-7306
- Volume :
- 3
- Issue :
- 9
- Database :
- MEDLINE
- Journal :
- Molecular and cellular biology
- Publication Type :
- Academic Journal
- Accession number :
- 6355826
- Full Text :
- https://doi.org/10.1128/mcb.3.9.1545-1551.1983