Enzyme-mediated dichloromethane toxicity and mutagenicity of bacterial and mammalian dichloromethane-active glutathione S-transferases.
The kinetic properties of bacterial and rat liver glutathione S-transferases (GST) active with dichloromethane (DCM) were compared. The theta class glutathione S-transferase (rGSTTI-1) from rat liver had an affinity for dihalomethanes lower by three orders of magnitude (K(app) > 50 mM) than the bacterial DCM dehalogenase/GST from Methylophilus sp. DM11. Unlike the bacterial DCM dehalogenase, the rat enzyme was unable to support growth of the dehalogenase minus Methylobacterium sp. DM4-2cr mutant with DCM. Moreover, the presence of DCM inhibited growth with methanol of the DM4-2cr transconjugant expressing the rat liver GSTT1-1. In Salmonella typhimurium TA1535, expression of rat and bacterial DCM-active GST from a plasmid in the presence of DCM yielded up to 5.3 times more reversions to histidine prototrophy in the transconjugant expressing the rat enzyme. Under the same conditions, however, GST-mediated conversion of DCM to formaldehyde was lower in cell-free extracts of the transconjugant expressing the rat GSTT1 than in the corresponding strain expressing the bacterial DCM dehalogenase. This provided new evidence that formaldehyde was not the main toxicant associated with GST-mediated DCM conversion, and indicated that an intermediate in the transformation of DCM by GST, presumably S-chloromethylglutathione, was responsible for the observed effects. The marked differences in substrate affinity of rat and bacterial DCM-active GST, as well as in the toxicity and genotoxicity associated with expression of these enzymes in bacteria, suggest that bacterial DCM dehalogenases/GST have evolved to minimise the toxic effects associated with glutathione-mediated catalysis of DCM conversion.
Gisi
D
D
Mikrobiologisches Institut, ETH Zürich, ETH Zentrum, Switzerland.
Leisinger
T
T
Vuilleumier
S
S
D000818:Animals; D001419:Bacteria / Q000201:enzymology*; D003001:Cloning, Molecular; D005557:Formaldehyde / Q000633:toxicity; D005982:Glutathione Transferase / Q000145:classification / Q000502:physiology*; D066298:In Vitro Techniques; D008099:Liver / Q000201:enzymology; D008752:Methylene Chloride / Q000378:metabolism / Q000633:toxicity*; D009152:Mutagenicity Tests; D009153:Mutagens / Q000633:toxicity*; D051381:Rats
D009153:Mutagens; D005557:Formaldehyde; D008752:Methylene Chloride; D005982:Glutathione Transferase