Dynamics of protein-protein interactions at the MscL periplasmic-lipid interface.
MscL, the highly conserved bacterial mechanosensitive channel of large conductance, is one of the best studied mechanosensors. It is a homopentameric channel that serves as a biological emergency release valve that prevents cell lysis from acute osmotic stress. We previously showed that the periplasmic region of the protein, particularly a single residue located at the TM1/periplasmic loop interface, F47 of Staphylococcus aureus and I49 of Escherichia coli MscL, plays a major role in both the open dwell time and mechanosensitivity of the channel. Here, we introduced cysteine mutations at these sites and found they formed disulfide bridges that decreased the channel open dwell time. By scanning a likely interacting domain, we also found that these sites could be disulfide trapped by addition of cysteine mutations in other locations within the periplasmic loop of MscL, and this also led to rapid channel kinetics. Together, the data suggest structural rearrangements and protein-protein interactions that occur within this region upon normal gating, and further suggest that locking portions of the channel into a transition state decreases the stability of the open state.
Department of Physiology, UT Southwestern Medical Center at Dallas, Dallas, Texas.
Dalian
D
Zhong
Department of Physiology, UT Southwestern Medical Center at Dallas, Dallas, Texas.
Li-Min
LM
Yang
Department of Physiology, UT Southwestern Medical Center at Dallas, Dallas, Texas. Electronic address: paul.blount@utsouthwestern.edu.
Paul
P
Blount
D000595:Amino Acid Sequence; D003545:Cysteine; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007473:Ion Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D050356:Lipid Metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009154:Mutation; D019897:Periplasm / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry; D055672:Static Electricity
D029968:Escherichia coli Proteins; D007473:Ion Channels; C086596:MscL protein, E coli; D021122:Protein Subunits; D003545:Cysteine