Effect of glutathione on aconitase in Escherichia coli.
The effect of glutathione (GSH) on the superoxide-sensitive [4Fe-4S]-containing aconitase of Escherichia coli was explored. A mutant deficient in GSH biosynthesis, designated gshA, grew slower in a defined medium than did the parental strain and this effect was more pronounced when succinate was supplied as the carbon source in place of glucose. This suggested that the citric acid cycle was compromised in the gshA strain. Aconitase activity was approximately 25% lower in GSH-deficient cells growing on either glucose or succinate, and was lower still in strains producing less superoxide dismutase. Addition of GSH to the medium stimulated growth of the gshA strain on succinate. It also elevated the aconitase activity in the presence of chloramphenicol, which was added to block protein synthesis. Dithiothreitol and 2-mercaptoethanol were much less effective in this regard than was GSH. Exposure of cultures to 4.2 atm O2 caused a rapid decline in aconitase activity and this was the case in both GSH-proficient and GSH-deficient E. coli; however, the reactivation which was seen when the hyperoxic exposure was terminated was significantly impaired in the gshA strain. There is a dynamic balance between inactivation of aconitase by superoxide and reactivation by Fe(II) and this balance is altered in GSH-deficient E. coli. GSH may facilitate reactivation of aconitase, and of other [4Fe-4S]-containing dehydratases, by increasing the rate of transfer of Fe(II) to the [3Fe-4S] site.
Gardner
P R
PR
Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710.
Fridovich
I
I
D000154:Aconitate Hydratase / Q000378:metabolism*; D004229:Dithiothreitol / Q000494:pharmacology; D004793:Enzyme Reactivators / Q000494:pharmacology; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology* / Q000254:growth & development; D005978:Glutathione / Q000235:genetics / Q000494:pharmacology*; D007700:Kinetics; D008623:Mercaptoethanol / Q000494:pharmacology; D009154:Mutation; D010100:Oxygen / Q000494:pharmacology; D013386:Succinates / Q000378:metabolism; D019802:Succinic Acid; D013482:Superoxide Dismutase / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism
D004793:Enzyme Reactivators; D013386:Succinates; D013481:Superoxides; D008623:Mercaptoethanol; D019802:Succinic Acid; D013482:Superoxide Dismutase; D000154:Aconitate Hydratase; D005978:Glutathione; D010100:Oxygen; D004229:Dithiothreitol