We recently found that budding yeast Saccharomyces cerevisiae sigma1278b, but not genome project strain S288C, has a gene conferring resistance to L-azetidine-2-carboxylic acid (AZC), a toxic four-membered ring analogue of L-proline. Also, the gene, designated as MPR1, encodes a novel acetyltransferase that detoxifies AZC via acetylation. We now report the results of subsequent work. On a homology search with MPR1, we detected a gene in fission yeast Schizosaccharomyces pombe. This gene, designated as ppr1(+) (pombe MPR1), is responsible for the AZC-resistance of S. pombe as judged from the results of gene disruption and overexpression experiments. Escherichia coli cells expressing ppr1(+), like ones expressing MPR1, were resistant to AZC and produced an AZC acetyltransferase. We further found that the enzymes encoded by MPR1 and ppr1(+) were homodimers, and catalyzed the acetylation of AZC but not any other L-proline-related compounds. Ppr1p was more thermostable than Mpr1p, although Ppr1p had a lower optimum temperature than Mpr1p. The higher AZC acetylation activity of Mpr1p, in comparison to that of Ppr1p, was attributed to the larger k(cat)/K(m) value for acetyl-CoA of Mpr1p than that of Ppr1p.