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 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.
Smith
D J
DJ
Glaxo Institute for Molecular Biology, Geneva, Switzerland.
Proudfoot
A E
AE
Detiani
M
M
Wells
T N
TN
Payton
M A
MA
D000595:Amino Acid Sequence; D001483:Base Sequence; D002176:Candida albicans / Q000235:genetics*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005800:Genes, Fungal; D008358:Mannose / Q000378:metabolism; D008359:Mannose-6-Phosphate Isomerase / Q000096:biosynthesis / Q000235:genetics*; D008969:Molecular Sequence Data
D008359:Mannose-6-Phosphate Isomerase; D008358:Mannose