The VIG9 gene products from the human pathogenic fungi Candida albicans and Candida glabrata encode GDP-mannose pyrophosphorylase.
We have identified two genomic DNA fragments from the human pathogenic fungi, Candida albicans (CaVIG9) and Candida glabrata (CgVIG9) that encode GDP-mannose pyrophosphorylase, a key enzyme for protein glycosylation. The VIG9 homologues of CaVIG9 and CgVIG9 complement an identified protein glycosylation-defective mutation, vig9, of Saccharomyces cerevisiae. The nucleotide sequences of the ORFs, which are 83 and 90% identical to that of the ScVIG9 protein, respectively, showed a predicted gene product homologous to S. cerevisiae GDP-mannose pyrophosphorylase. We examined the enzyme activity of a glutathione S-transferase fusion of each VIG9 gene to synthesize GDP mannose in the cell extracts of a heterologous Escherichia coli expression system. We also developed a method for detecting the enzyme activity using a non-radioactive substrate that would be applicable to high throughput screening.
Ohta
A
A
Department of Mycology, Nippon Roche Research Center, Kamakura, Kanagawa, Japan.
Chibana
H
H
Arisawa
M
M
Sudoh
M
M
D000595:Amino Acid Sequence; D002176:Candida albicans / Q000235:genetics*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D005816:Genetic Complementation Test; D006155:Guanosine Diphosphate Mannose / Q000032:analysis / Q000096:biosynthesis; D008969:Molecular Sequence Data; D009713:Nucleotidyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D005656:Fungal Proteins; D006155:Guanosine Diphosphate Mannose; D009713:Nucleotidyltransferases; C026917:mannose 1-phosphate guanylyltransferase