Hydrophilicity of a Single Residue within MscL Correlates with Increased Channel Mechanosensitivity

AbstractMechanosensitive channel large (MscL) encodes the large conductance mechanosensitive channel of the Escherichia coli inner membrane that protects bacteria from lysis upon osmotic shock. To elucidate the molecular mechanism of MscL gating, we have comprehensively substituted Gly22 with all other common amino acids. Gly22 was highlighted in random mutagenesis screens of E. coli MscL (Ou et al., 1998, Proc. Nat. Acad. Sci. USA. 95:11471–11475). By analogy to the recently published MscL structure from Mycobacterium tuberculosis (Chang et al., 1998, Science. 282:2220–2226), Gly22 is buried within the constriction that closes the pore. Substituting Gly22 with hydrophilic residues decreased the threshold pressure at which channels opened and uncovered an intermediate subconducting state. In contrast, hydrophobic substitutions increased the threshold pressure. Although hydrophobic substitutions had no effect on growth, similar to the effect of an MscL deletion, channel hyperactivity caused by hydrophilic substitutions corrrelated with decreased proliferation. These results suggest a model for gating in which Gly22 moves from a hydrophobic, and through a hydrophilic, environmment upon transition from the closed to open conformation