Domain organization of the MscS mechanosensitive channel of Escherichia coli.
The major structural features of the Escherichia coli MscS mechanosensitive channel protein have been explored using alkaline phosphatase (PhoA) fusions, precise deletions and site-directed mutations. PhoA protein fusion data, combined with the positive-inside rule, strongly support a model in which MscS crosses the membrane three times, adopting an N(out)-C(in) configuration. Deletion data suggest that the C-terminal domain of the protein is essential for the stability of the MscS channel, whereas the protein will tolerate small deletions at the N-terminus. Four mutants that exhibit either gain-of-function (GOF) or loss-of-function have been identified: a double mutation I48D/S49P inactivates MscS, whereas the MscS mutants T93R, A102P and L109S cause a strong GOF phenotype. The similarity of MscS to the last two domains of MscK (formerly KefA) is reinforced by the demonstration that expression of a truncated MscK protein can substitute for MscL and MscS in downshock survival assays. The data derived from studies of the organization, conservation and the influence of mutations provide significant insights into the structure of the MscS channel.
Miller
Samantha
S
Department of Molecular and Cell Biology, University of Aberdeen, Institute of Medical Sciences, Aberdeen AB25 2ZD, UK.
Bartlett
Wendy
W
Chandrasekaran
Subramanian
S
Simpson
Sally
S
Edwards
Michelle
M
Booth
Ian R
IR
D000469:Alkaline Phosphatase / Q000737:chemistry / Q000378:metabolism; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001483:Base Sequence; D002462:Cell Membrane / Q000201:enzymology; D017931:DNA Primers; D004926:Escherichia coli / Q000502:physiology*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D007473:Ion Channels / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D015183:Restriction Mapping; D012680:Sensitivity and Specificity; D017384:Sequence Deletion
D017931:DNA Primers; D029968:Escherichia coli Proteins; D007473:Ion Channels; C469542:MscS protein, E coli; D000469:Alkaline Phosphatase