CLONING AND HETEROLOGOUS EXPRESSION OF THE CANDIDA-ALBICANS GENE PMI-1 ENCODING PHOSPHOMANNOSE ISOMERASE

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 PMI1 gene. This gene, which is unique in the C. albicans genome, can functionally complement PMI-deficient mutants of both S. cerevisiae and Escherichia cell. The DNA sequence of the PMI1 gene predicts a protein with 64.1% identity to PMI from S. cerevisiae. Sequential gene disruption of PIMI1 produces a strain with an auxotrophic requirement for D-mannose. The heterologous expression of the PMI1 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.