Structure of the MscL Homolog from Mycobacterium tuberculosis : A Gated Mechanosensitive Ion Channel

Abstract

Mechanosensitive ion channels play a critical role in transducing physical stresses at the cell membrane into an electrochemical response. The MscL family of large-conductance mechanosensitive channels is widely distributed among prokaryotes and may participate in the regulation of osmotic pressure changes within the cell. In an effort to better understand the structural basis for the function of these channels, the structure of the MscL homolog from Mycobacterium tuberculosis was determined by x-ray crystallography to 3.5 angstroms resolution. This channel is organized as a homopentamer, with each subunit containing two transmembrane α helices and a third cytoplasmic α helix. From the extracellular side, a water-filled opening approximately 18 angstroms in diameter leads into a pore lined with hydrophilic residues which narrows at the cytoplasmic side to an occluded hydrophobic apex that may act as the channel gate. This structure may serve as a model for other mechanosensitive channels, as well as the broader class of pentameric ligand-gated ion channels exemplified by the nicotinic acetylcholine receptor.

Structural basis of water-specific transport through the AQP1 water channel

“…Extension of phases and model refinement to 2.2 Å was accomplished with a data set obtained from crystals grown in the presence of gold cyanide but for which no multi-wavelength anomalous differences were observed. Others have also obtained improvements in crystal order using gold cyanide as a crystallization additive 33 . While the diffraction data from these crystals extended to 2.1 Å it was significantly anisotropic and resulted in a reduction in the percentage of statistically reliable data in the direction of the h, k plane.…”

Spectator no more, the role of the membrane in regulating ion channel function

“…The activation of MscL in droplet interface bilayers [17] may herald novel nano-technological applications of the protein. The closed crystal structure of pentameric M. tuberculosis MscL [4] has been known for several years. Native mass spectrometry of MscL suggested the in vitro oligomerisation state (pentamer vs tetramer) is influenced by temperature, detergent and amino acid variability as well as by the precise construct used [18].…”

“…The "horizontal" helix has no conservation in sequence or length (Fig 3). This helix radiates outwards (almost 90 0 ) to the central pore in MscS[34], MscL [4] and TRPV1 [38] channels. In TRAAK [8] an acyl chain from a lipid obstructs the ion conduction pathway (Fig 3C).…”

“…A change in the perception of lipids as actively regulating protein function came from the study of a pair of bacterial mechanosensitive channels, the heptameric mechanosensing channel of small conductance (MscS) [2,3] and the pentameric mechanosensing channel of large conductance (MscL) [4,5]. These proteins are controlled by changes in the lipid bilayer that arise from increased pressure inside the cell (known as turgor).…”

An approach to crystallizing proteins by metal‐mediated synthetic symmetrization

“…Although high-throughput robotics now makes it possible to screen vast numbers of crystallization conditions, some proteins remain recalcitrant to crystallization. Many approaches have been developed to improve the success rate for crystallization, for example, systematically truncating the target protein, 1,2 methylating the lysine residues, 3,4 removing post-translational modifications, [5][6][7] screening homologues of the target protein for crystallization, 8,9 fusing the target protein to a carrier protein, [10][11][12][13] crystallizing racemic mixtures of the target protein, [14][15][16][17][18][19][20] and cocrystallizing the target protein with antibodies or other binding proteins. [21][22][23] A number of those methods have been reviewed.…”