Abstract
k9 killer toxin from Hansenula mrakii was used to select a number of resistant mutants from Saccharomyces cerevisiae. Preliminary biochemical and genetic studies showed that some of them acquired structural defects in the cell wall. One of these mutants, the knr4-1 mutant, displays a number of cell wall defects, including osmotic sensitivity; sensitivity to cercosporamide, a known antifungal agent; and resistance to Zymolyase, a (1,3)-beta-glucanase. We report here the isolation and analysis of the KNR4 gene. DNA sequence analysis revealed an uninterrupted open reading frame which contains five potential start codons. The longest coding template encodes a protein of 505 amino acids with a calculated molecular mass of 57,044 Da. A data base search revealed 100% identity with a nuclear protein, SMI1p. Disruption of the KNR4 locus does not result in cell death; however, it leads to reduced levels of both (1,3)-beta-glucan synthase activity and (1,3)-beta-glucan content in the cell wall. The gene was mapped to the right arm of chromosome VII.
MeSH terms
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Amino Acid Sequence
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Base Sequence
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Benzofurans / toxicity
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Blotting, Western
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Chromosomes, Fungal
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Cloning, Molecular / methods
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DNA, Fungal / isolation & purification
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DNA, Fungal / metabolism
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Drug Resistance, Microbial / genetics*
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Fungal Proteins / biosynthesis*
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Fungal Proteins / genetics
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Fungal Proteins / isolation & purification
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Genes, Fungal*
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Glucans / biosynthesis*
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Glucosyltransferases / biosynthesis*
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Glucosyltransferases / metabolism
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Hydrolases / toxicity
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Killer Factors, Yeast
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Kinetics
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Membrane Proteins*
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Molecular Sequence Data
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Molecular Weight
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Open Reading Frames
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Pichia / metabolism
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Protein Biosynthesis
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Proteins / toxicity
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Recombinant Proteins / biosynthesis
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Recombinant Proteins / isolation & purification
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Restriction Mapping
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Saccharomyces cerevisiae / drug effects
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Saccharomyces cerevisiae / genetics*
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Saccharomyces cerevisiae / metabolism*
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Saccharomyces cerevisiae Proteins*
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Schizosaccharomyces pombe Proteins*
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Sequence Homology, Amino Acid
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Transcription Factors
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beta-Glucans*
Substances
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Benzofurans
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DNA, Fungal
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Fungal Proteins
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Glucans
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Killer Factors, Yeast
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Membrane Proteins
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Proteins
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Recombinant Proteins
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SMI1 protein, S cerevisiae
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Saccharomyces cerevisiae Proteins
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Schizosaccharomyces pombe Proteins
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Transcription Factors
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beta-Glucans
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cercosporamide
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zymolyase
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beta-1,3-glucan
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Glucosyltransferases
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1,3-beta-glucan synthase
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Hydrolases