C termini of the Escherichia coli mechanosensitive ion channel (MscS) move apart upon the channel opening.
Heptameric YggB is a mechanosensitive ion channel (MscS) from the inner membrane of Escherichia coli. We demonstrate, using the patch clamp technique, that cross-linking of the YggB C termini led to irreversible inhibition of the channel activities. Application of Ni(2+) to the YggB-His(6) channels with the hexahistidine tags added to the ends of their C termini also resulted in a marked but reversible decrease of activities. Western blot revealed that YggB-His(6) oligomers are more stable in the presence of Ni(2+), providing evidence that Ni(2+) is coordinated between C termini from different subunits of the channel. Intersubunit coordination of Ni(2+) affecting channel activities occurred in the channel closed conformation and not in the open state. This may suggest that the C termini move apart upon channel opening and are involved in the channel activation. We propose that the as yet undefined C-terminal region may form a cytoplasmic gate of the channel. The results are discussed and interpreted based on the recently released quaternary structure of the channel.
Koprowski
Piotr
P
Department of Cell Biology, Nencki Institute of Experimental Biology; 3, Pasteur Street, 02-093 Warsaw, Poland.
Kubalski
Andrzej
A
D000595:Amino Acid Sequence; D001483:Base Sequence; D017931:DNA Primers; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism*; D015640:Ion Channel Gating; D007473:Ion Channels / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009532:Nickel / Q000737:chemistry; D018408:Patch-Clamp Techniques
D017931:DNA Primers; D029968:Escherichia coli Proteins; D007473:Ion Channels; C469542:MscS protein, E coli; D009532:Nickel