The allantoinase (DAL1) gene from Saccharomyces cerevisiae has been cloned, sequenced, and found to encode a 472 amino acid protein with a Mr of 52,028. DAL1 is expressed in an inducer-independent manner in strain M970 (sigma 1278b genetic background) and modestly responds to mutation of the dal80 locus. Expression was also sensitive to nitrogen catabolite repression (NCR). Correlated with these expression characteristics, the upstream region of DAL1 contained five copies of a sequence that is homologous to the DAL UASNTR element previously shown to be required for transcriptional activation and NCR sensitivity of the DAL5 and DAL7 genes. Missing from the DAL1 5' flanking region were any sequences with significant homology to the DAL7 UIS element required for response to inducer. These observations further support the roles of UASNTR and DAL7 UIS in the regulation of allantoin pathway gene expression.