Abstract: Most recently， a GSHindependent glyoxalase pathway was identified in Escherichia coli， Caenorhabditis elegans， mice and humans. Mechanisms of MG detoxicification include the glutathione (GSH)dependent pathway consisting of glyoxalase Ⅰ (GLO1) and glyoxalase Ⅱ (GLO2)， and GSHindependent pathway catalyzed by glyoxalase Ⅲ (GLO3). E. coli Hsp31 and metazoan DJ1 proteins， belong to two different subfamilies of the Hsp31/DJ1 superfamily， have been demonstrated to have GLO3 activity. However， genes encoding GLO3 have not been reported in fungi. We performed a systematic survey of homologs of Hsp31 and DJ1 in fungi. We found that Hsp31 proteins are widely distributed among different fungal groups， whereas DJ1 proteins have a very limited distribution in fungi. Phylogenetic analysis of these sequences revealed that fungal Hsp31 and DJ1 proteins form two distinct monophyletic groups， and that fungal Hsp31 proteins can clearly be divided into different classes. One Saccharomyces cerevisiae Hsp31 protein (ScHsp31) and Schizosaccharomyces pombe DJ1 (SpDJ1) have GLO3 activity in vitro. Our results revealed that fungal homologs of the Hsp31/DJ1 superfamily could be GLO3， which may have some roles in protecting cells from MG toxicity. Our results also support the view that GLO3 activity likely evolved through convergent evolution.

Key words: fungi, methylglyoxal, GLO3, convergent evolution, HPLC