Exploring the pH Sensitivity of E. Coli MscS

Abstract

The mechanosensitive channel of small conductance (MscS) from Escherichia coli (E. coli) gates in response to mechanical tension that is generated from hypo-osmotic shock. The gating mechanism of E. coli MscS, in response to mechanical tension, is similar to that of a Jack-in-the-Box, the channel springs into the open state when the applied extrinsic tension overcomes the intrinsic lateral tension of the membrane. E. coli MscS is the founding member of the MscS superfamily, a superfamily of 15 subfamilies that all contain significant homology to the pore lining helix of E. coli MscS. In the early identification of MscS a rapid desensitization of MscS when pressure is applied at low pH is observed, however when patched at physiological pH E. coli MscS does not desensitize. To identify the region of the channel that evokes pH sensitivity we will use asymmetric and symmetric pH patch buffer. A recent study explored how significant alterations to the most conserved residues impacted E. coli MscS function, one residue F127 showed significant alteration to the gating phenotype when mutated to different residues. The results of the previous study show that the rapid desensitization classically observed in low pH are observed in a F127 mutations. This suggests that F127 is involved in a long-range interaction, potentially a cation-π with K60. Based on the location of this interaction in multiple crystal structures, the cation-π is potentially involved in the desensitized state of the channel. Through patch clamp electrophysiology we will understand the role of a pH sensitive region in the gating cycle of E. coli MscS.