Cloning and sequence analysis of the Candida tropicalis URA3 gene encoding orotidine-5'-phosphate decarboxylase.
An integrative transformation system was established for a phenol-utilizing strain of Candida tropicalis M4. The system is based on an auxotrophic mutant host of C. tropicalis U-6 that is defective in orotidine-5'-phosphate decarboxylase (ODCase). As a selectable marker, we isolated and characterized the C. tropicalis URA3 gene, which codes for ODCase. The gene was cloned by complementation of the ura3 mutation of Sachharomyces cerevisiae SHY-3 and the pyrF mutation of Escherichia coli. The C. tropicalis U-6 was transformed by plasmid containing the C. tropicalis URA3 gene at a frequency of 1 to 10 transformants per microgram of plasmid DNA. When the URA3 gene was expressed in E. coli minicells, a 30-kDa protein was identified. Nucleotide sequence analysis revealed the presence of an open reading frame, encoding a protein of 268 amino acids with a calculated molecular mass of 29.7 kDa. The nucleotide sequence of URA3 gene and its deduced amino acid sequence showed significant homology to those of the ODCase of other fungal species.
Su
J H
JH
Department of Industrial Safety and Hygiene, Chia-Nan College of Pharmacy and Science, Tainan, Taiwan, Republic of China.
Hsia
J H
JH
Chang
M C
MC
D001483:Base Sequence; D002175:Candida / Q000201:enzymology* / Q000235:genetics*; D003001:Cloning, Molecular; D005800:Genes, Fungal / Q000235:genetics; D008969:Molecular Sequence Data; D009154:Mutation; D016366:Open Reading Frames / Q000235:genetics; D009964:Orotidine-5'-Phosphate Decarboxylase / Q000235:genetics*; D019800:Phenol / Q000378:metabolism; D011994:Recombinant Proteins / Q000032:analysis; D012441:Saccharomyces cerevisiae / Q000235:genetics
D011994:Recombinant Proteins; D019800:Phenol; D009964:Orotidine-5'-Phosphate Decarboxylase