The orotidine-5′-phosphate decarboxylase gene (URA3) of a methylotrophic yeast, Candida boidinii: Nucleotide sequence and its expression in Escherichia coli

Abstract

Abstract Previously, we cloned a DNA fragment from a genomic library of a methylotrophic yeast, Candida boidinii . This 3.5-kb Sal I fragment was capable of complementing the pyrF mutation in Escherichia coli . In this report, we identify this fragment as that harboring an orotidine-5′-phosphate decarboxylase (ODCase) gene ( C. boidinii URA3 ); we have also determined the complete DNA sequence of the C. boidinii URA3 gene. The deduced amino acid sequence of the gene showed homology to ODCase genes from other sources, and it could complement the ura3 mutation of Saccharomyces cerevisiae . The DNA fragment, which harbored the C. boidinii URA3 gene, was able to express ODCase activity in the E. coli pyrF mutant strain without an exogenous E. coli promoter. From nested-deletion analysis, both the 5′-(136 bp) and 3′-(58 bp) flanking regions were shown to be required for pyrF -complementation of the E. coli mutant. The 5′-flanking region had sequences homologous to E. coli promoter consensus sequences (−35 and −10 regions) which may function in the expression of the C. boidinii URA3 gene in E. coli .