Final Report Summary - MECHANOSENSATION (What is the molecular mechanism of mechanosensation? Mechanosensitive channel of large conductance, MscL, as a model) The mechanosensitive ion channels are molecular transducers of mechanical force in all living cells. The best-characterized mechanosensitive ion channel is mechanosensitive channel of large conductance (MscL) from Escherichia coli. It senses membrane tension invoked by sudden hypoosmotic stress and acts as an emergency valve by opening a large, transient, aqueous pore in the membrane. Despite the importance of these channels and ongoing efforts to explain their functioning, the molecular mechanism of MS channel gating remains elusive and controversial. To understand the gating mechanism of MscL, we developed a method that allows single-subunit resolution for manipulating and monitoring of homo-oligomeric proteins. With that, we demonstrated experimentally the hydrophobic and asymmetric gating of MscL and how the channel opening and closing are initiated. We defined the pore diameter of this ion channel at various stages of its gating and showed that in the absence of applied tension, the hydrophobic gating alone is not enough to open the channel fully. In addition, by using individual heteropentamers in ion-mobility-mass spectrometry, we could observe the global structural changes of MscL during its gating, for the first time. Recently, we had control over the channel gating by interfering with its interaction with the lipid bilayer in a reversible manner and showed that MscL from different homologues reacts differently to the global and local changes in the lipid bilayer properties. We developed a solvatometric probe and followed the microenvironment of pore residues and could observe very early movements of the pore. Since MscL converts the mechanical forces on the membrane directly into such structural changes, this latest finding will pave the way to model the mechanical forces acting on the protein. MscL is an exciting tool also for nanotechnology: its large, nonselective pore allows the passage of even small peptides; it does not require cellular components for its function and remains functional in synthetic lipid environments. By exploring its hydrophobic gating, we converted MscL into a light- and pH-sensory valve in drug delivery liposomes. Recently, by using magnetic resonance spectroscopy and imaging, we showed that MscL-functionalized liposomes are sensitive enough to detect the mildly acidic pH conditions of the tumor microenvironment and release their drug into C6 glioma tumors, in vivo. Finally, we demonstrated the potential of MscL as a sensory component on Si/SiO2 chips.
