A gene whose product enhances glyoxalase I activity in yeast Saccharomyces cerevisiae cells was cloned as a 6-kbp DNA fragment in the vector YEp13. The DNA fragment was subcloned in approximately 2 kbp and its base sequence determined. An open reading frame with 318 nucleotide pairs (encoding 106 amino acids) was found. According to Northern blotting analysis, the yeast transformed with the hybrid plasmid harboring the 2-kbp fragment yielded two different transcripts. From the DNA sequence of the fragment, it is deduced that the gene product would contain seven cysteine residues, five of which would be located near the carboxy terminus of the peptide.