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Membrane topology and multimeric structure of a mechanosensitive channel protein of Escherichia coli.
Author(s) -
Blount P.,
Sukharev S. I.,
Moe P. C.,
Schroeder M. J.,
Guy H. R.,
Kung C.
Publication year - 1996
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1002/j.1460-2075.1996.tb00860.x
Subject(s) - biology , mechanosensitive channels , escherichia coli , escherichia coli proteins , membrane protein , microbiology and biotechnology , biophysics , membrane , topology (electrical circuits) , ion channel , genetics , gene , receptor , mathematics , combinatorics
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.