Transcriptional and maturational effects of manganese and iron on the biosynthesis of manganese-superoxide dismutase in Escherichia coli.
Anaerobically grown Escherichia coli contain an enzymatically active iron superoxide dismutase (Fe2-FeSOD) and an inactive iron-substituted manganese superoxide dismutase (Fe2-MnSOD). The anaerobic electron sink, nitrate plus paraquat, enhanced biosynthesis of the MnSOD polypeptide, with accumulation of inactive Fe2-MnSOD. The oxidant, diamide, in contrast, allowed anaerobic production of the active forms of MnSOD, i.e. Mn2-MnSOD and Mn/Fe-MnSOD. Nutritional supplementation with Mn(II) favored occupancy of the MnSOD active site with manganese and allowed anaerobic accumulation of Mn2-MnSOD in the absence of diamide. Enrichment of the anaerobic growth medium with Fe(II) both suppressed biosynthesis of the MnSOD polypeptide and inhibited formation of the active manganese-containing forms. A tac-sodA operon fusion was used to examine the effects of chelating agents and metals on maturation of nascent MnSOD, independent from the transcriptional effects these agents impose. Isopropyl-1-thio-beta-D-galactopyranoside (IPTG) elicited anaerobic biosynthesis of MnSOD, which accumulated as the inactive Fe2-MnSOD. Diamide, with IPTG, allowed formation of active Mn/Fe-MnSOD while 1,10-phenanthroline with IPTG resulted in accumulation of Mn2-MnSOD. These results suggest that iron participates in the redox-sensitive control of the formation of active MnSOD at two levels, i.e. that of transcription as well as that of maturation. During maturation of the nascent MnSOD polypeptide, iron and manganese compete for the metal-binding site; anaerobic conditions favor iron-binding, whereas oxidants, such as dioxygen or diamide, favor binding of manganese.
Privalle
C T
CT
Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710.
Fridovich
I
I
D015153:Blotting, Western; D003958:Diamide / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology* / Q000378:metabolism; D007501:Iron / Q000378:metabolism*; D008345:Manganese / Q000378:metabolism*; D009876:Operon; D010618:Phenanthrolines / Q000494:pharmacology; D013482:Superoxide Dismutase / Q000096:biosynthesis* / Q000378:metabolism; D014158:Transcription, Genetic
D010618:Phenanthrolines; D003958:Diamide; D008345:Manganese; D007501:Iron; D013482:Superoxide Dismutase; C025205:1,10-phenanthroline