Activation of bacterial channel MscL in mechanically stimulated droplet interface bilayers.
MscL, a stretch-activated channel, saves bacteria experiencing hypo-osmotic shocks from lysis. Its high conductance and controllable activation makes it a strong candidate to serve as a transducer in stimuli-responsive biomolecular materials. Droplet interface bilayers (DIBs), flexible insulating scaffolds for such materials, can be used as a new platform for incorporation and activation of MscL. Here, we report the first reconstitution and activation of the low-threshold V23T mutant of MscL in a DIB as a response to axial compressions of the droplets. Gating occurs near maximum compression of both droplets where tension in the membrane is maximal. The observed 0.1-3 nS conductance levels correspond to the V23T-MscL sub-conductive and fully open states recorded in native bacterial membranes or liposomes. Geometrical analysis of droplets during compression indicates that both contact angle and total area of the water-oil interfaces contribute to the generation of tension in the bilayer. The measured expansion of the interfaces by 2.5% is predicted to generate a 4-6 mN/m tension in the bilayer, just sufficient for gating. This work clarifies the principles of interconversion between bulk and surface forces in the DIB, facilitates the measurements of fundamental membrane properties, and improves our understanding of MscL response to membrane tension.
Department of Mechanical Engineering, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061, United States.
Joseph S
JS
Najem
School of Biomedical Engineering and Sciences, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061, United States.
Myles D
MD
Dunlap
Department of Biology, University of Maryland, College Park, Maryland 20742, United States.
Ian D
ID
Rowe
College of Engineering, University of Georgia, Athens, Georgia 30602, United States.
Eric C
EC
Freeman
Department of Engineering Science and Mechanics, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061, United States.
John W
JW
Grant
Department of Biology, University of Maryland, College Park, Maryland 20742, United States.
Sergei
S
Sukharev
College of Engineering, University of Georgia, Athens, Georgia 30602, United States.
Donald J
DJ
Leo
D000200:Action Potentials; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D007473:Ion Channels / Q000235:genetics / Q000378:metabolism*; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism*; D055595:Mechanical Phenomena; D009154:Mutation; D013500:Surface Tension
D029968:Escherichia coli Proteins; D007473:Ion Channels; D008051:Lipid Bilayers; C086596:MscL protein, E coli