Structural Investigation of MscL Gating Using Experimental Data and Coarse Grained MD Simulations

The mechanosensitive channel of large conductance (MscL) has become a model system in which to understand mechanosensation, a process involved in osmoregulation and many other physiological functions. While a high resolution closed state structure is available, details of the open structure and the gating mechanism remain unknown. In this study we combine coarse grained simulations with restraints from EPR and FRET experiments to study the structural changes involved in gating with much greater level of conformational sampling than has previously been possible. We generated a set of plausible open pore structures that agree well with existing open pore structures and gating models. Most interestingly, we found that membrane thinning induces a kink in the upper part of TM1 that causes an outward motion of the periplasmic loop away from the pore centre. This previously unobserved structural change might present a new mechanism of tension sensing and might be related to a functional role in osmoregulation.The study was supported by a grant from the Australian Research Council. The simulations were carried out using computer time from iVEC and a Merit Allocation Scheme on the NCI National Facility at the Australian National University. ED was supported by a Jean Rogerson Postgraduate scholarship and the Beryl Henderson Memorial Grant by the Australian Federation of University Women ACT. Websites of funding agencies: http://nci.org.au/access/merit-allocationscheme/, http://www.ivec.org/ http://www.arc.gov.au/ncgp/default.htm, http://spe.publishing.uwa.edu.au/latest/scholarships/postgraduate/rogerson, http://www. afgw.org.au/what-we-do/scholarships-2/ The authors hereby confirm that the funding agencies had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Bacterial mechanosensitive channels : progress towards an understanding of their roles in cell physiology

Open Access funded by Wellcome Trust Under a Creative Commons license Thanks to all members of the Aberdeen group, collaborators and friends whose discussions have spurred the development of the MS channel field. Special thanks to Doug Rees, Diane Newman and Rob Phillips for their support and hospitality at Caltech. Unique insights have been provided by members of the Newman and Phillips research groups, particularly, Caj Neubauer, Gargi Kulkarni and Megan Bergkessel, Heun Jin Lee and Maja Bialecka-Fornal. The author's research on MS channels is supported by a grant from The Wellcome Trust (WT092552MA) and the BBSRC (BB/H017917/1). The author is a Leverhulme Emeritus Fellow and this work was supported in part by a CEMI Visiting Faculty Fellowship from Caltech.Peer reviewedPublisher PD

Analytic models for mechanotransduction: gating a mechanosensitive channel

Analytic estimates for the forces and free energy generated by bilayer deformation reveal a compelling and intuitive model for MscL channel gating analogous to the nucleation of a second phase. We argue that the competition between hydrophobic mismatch and tension results in a surprisingly rich story which can provide both a quantitative comparison to measurements of opening tension for MscL when reconstituted in bilayers of different thickness and qualitative insights into the function of the MscL channel and other transmembrane proteins

Hydrophobic gating of mechanosensitive channel of large conductance evidenced by single-subunit resolution

Mechanosensitive (MS) ion channels are membrane proteins that detect and respond to membrane tension in all branches of life. In bacteria, MS channels prevent cells from lysing upon sudden hypoosmotic shock by opening and releasing solutes and water. Despite the importance of MS channels and ongoing efforts to explain their functioning, the molecular mechanism of MS channel gating remains elusive and controversial. Here we report a method that allows single-subunit resolution for manipulating and monitoring “mechanosensitive channel of large conductance” from Escherichia coli. We gradually changed the hydrophobicity of the pore constriction in this homopentameric protein by modifying a critical pore residue one subunit at a time. Our experimental results suggest that both channel opening and closing are initiated by the transmembrane 1 helix of a single subunit and that the participation of each of the five identical subunits in the structural transitions between the closed and open states is asymmetrical. Such a minimal change in the pore environment seems ideal for a fast and energy-efficient response to changes in the membrane tension.

Quantitative description of ion transport via plasma membrane of yeast and small cells

Modeling of ion transport via plasma membrane needs identification and quantitative understanding of the involved processes. Brief characterization of main ion transport systems of a yeast cell (Pma1, Ena1, TOK1, Nha1, Trk1, Trk2, non-selective cation conductance) and determining the exact number of molecules of each transporter per a typical cell allow us to predict the corresponding ion flows. In this review a comparison of ion transport in small yeast cell and several animal cell types is provided. The importance of cell volume to surface ratio is emphasized. The role of cell wall and lipid rafts is discussed in respect to required increase in spatial and temporal resolution of measurements. Conclusions are formulated to describe specific features of ion transport in a yeast cell. Potential directions of future research are outlined based on the assumptions.Comment: 22 pages, 6 figures, 1 tabl