Iron is a ligand of SecA-like metal-binding domains in vivo

Tamar Cranford-Smith, Mohammed Jamshad, Mark Jeeves, Rachael A. Chandler, Jack Yule, Ashley Robinson, Farhana Alam, Karl A. Dunne, Edwin H. Aponte Angarita, Mashael Alanazi, Cailean Carter, Ian R. Henderson, Janet E. Lovett, Peter Winn, Timothy Knowles, Damon Huber

Research output: Contribution to journalArticlepeer-review

5 Citations (Scopus)
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Abstract

The ATPase SecA is an essential component of the bacterial Sec machinery, which transports proteins across the cytoplasmic membrane. Most SecA proteins contain a long C-terminal tail (CTT). In Escherichia coli, the CTT contains a structurally flexible linker domain and a small metal-binding domain (MBD). The MBD coordinates zinc via a conserved cysteine-containing motif and binds to SecB and ribosomes. In this study, we screened a high-density transposon library for mutants that affect the susceptibility of E. coli to sodium azide, which inhibits SecA-mediated translocation. Results from sequencing this library suggested that mutations removing the CTT make E. coli less susceptible to sodium azide at subinhibitory concentrations. Copurification experiments suggested that the MBD binds to iron and that azide disrupts iron binding. Azide also disrupted binding of SecA to membranes. Two other E. coli proteins that contain SecA-like MBDs, YecA and YchJ, also copurified with iron, and NMR spectroscopy experiments indicated that YecA binds iron via its MBD. Competition experiments and equilibrium binding measurements indicated that the SecA MBD binds preferentially to iron and that a conserved serine is required for this specificity. Finally, structural modelling suggested a plausible model for the octahedral coordination of iron. Taken together, our results suggest that SecA-like MBDs likely bind to iron in vivo.
Original languageEnglish
JournalJournal of Biological Chemistry
VolumePapers in Press
Early online date2 Apr 2020
DOIs
Publication statusE-pub ahead of print - 2 Apr 2020

Keywords

  • Sec pathway
  • Iron
  • Protein secretion
  • Protein structure
  • Protein translocation
  • Metal ion-protein interaction

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