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Cloning and heterologous expression of the Candida albicans gene PMI 1 encoding phosphomannose isomerase
Author(s) -
Smith David J.,
Proudfoot Amanda E. I.,
Detiani Mariastella,
Wells Timothy N. C.,
Payton Mark A.
Publication year - 1995
Publication title -
yeast
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.923
H-Index - 102
eISSN - 1097-0061
pISSN - 0749-503X
DOI - 10.1002/yea.320110402
Subject(s) - biology , candida albicans , gene , heterologous expression , saccharomyces cerevisiae , microbiology and biotechnology , dna , genomic dna , accession number (library science) , corpus albicans , mutant , genetics , biochemistry , recombinant dna , genbank
Using a DNA fragment derived from the Saccharomyces cerevisiae phosphomannose isomerase (PMI) structural gene as a probe against a random ordered array library of genomic DNA from the pathogenic fungus Candida albicans , we have cloned the C. albicans PMI 1 gene. This gene, which is unique in the C. albicans genome, can functionally complement PMI‐deficient mutants of both S. cerevisiae and Escherichia coli . The DNA sequence of the PMI 1 gene predicts a protein with 64·1% identity to PMI from S. cerevisiae . Sequential gene disruption of PMI 1 produces a strain with an auxotrophic requirement for D‐mannose. The heterologous expression of the PMI 1 gene at levels up to 45% of total cell protein in E. coli leads to partitioning of the enzyme between the soluble and particulate fractions. The protein produced in the soluble fraction is indistinguishable in kinetic properties from the material isolated from C. albicans cells. The nucleotide sequence data reported here will appear in the EMBL database under Accession Number X82024.