Membrane topology and multimeric structure of a mechanosensitive channel protein of Escherichia coli

Paul Blount, Sergei I. Sukharev, Paul C. Moe, Matthew J. Schroeder, H. Robert Guy, Ching Kung

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Abstract

We have studied the membrane topology and multimeric structure of a mechanosensitive channel, MscL, which we previously isolated and cloned from Escherichia coli. We have localized this 15-kDa protein to the inner membrane and, by PhoA fusion, have shown that it contains two transmembrane domains with both the amino and carboxyl termini on the cytoplasmic side. Mutation of the glutamate at position 56 to histidine led to changes in channel kinetics which were dependent upon the pH on the periplasmic, but not cytoplasmic side of the membrane, providing additional evidence for the periplasmic positioning of this part of the molecule. Tandems of two MscL subunits expressed as a single polypeptide formed functional channels, suggesting an even number of transmembrane domains per subunit (amino and carboxyl termini on the same side of the membrane), and an even number of subunits per functional complex. Finally, cross-linking studies suggest that the functional MscL complex is a homohexamer. In summary, these data are all consistent with a protein domain assignment and topological model which we propose and discuss.

Original languageEnglish (US)
Pages (from-to)4798-4805
Number of pages8
JournalEMBO Journal
Volume15
Issue number18
StatePublished - Sep 16 1996

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Keywords

  • Mechanosensation
  • Mechanosensitive channel
  • MscL
  • Osmotic forces
  • Stretch activated channel

ASJC Scopus subject areas

  • Neuroscience(all)
  • Molecular Biology
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

Cite this

Blount, P., Sukharev, S. I., Moe, P. C., Schroeder, M. J., Guy, H. R., & Kung, C. (1996). Membrane topology and multimeric structure of a mechanosensitive channel protein of Escherichia coli. EMBO Journal, 15(18), 4798-4805.