Single residue substitutions that change the gating properties of a mechanosensitive channel in Escherichia coli.

MscL is a channel that opens a large pore in the Escherichia coli cytoplasmic membrane in response to mechanical stress. Previously, we highly enriched the MscL protein by using patch clamp as a functional assay and cloned the corresponding gene. The predicted protein contains a largely hydrophobic core spanning two-thirds of the molecule and a more hydrophilic carboxyl terminal tail. Because MscL had no homology to characterized proteins, it was impossible to predict functional regions of the protein by simple inspection. Here, by mutagenesis, we have searched for functionally important regions of this molecule. We show that a short deletion from the amino terminus (3 amino acids), and a larger deletion of 27 amino acids from the carboxyl terminus of this protein, had little if any effect in channel properties. We have thus narrowed the search of the core mechanosensitive mechanism to 106 residues of this 136-amino acid protein. In contrast, single residue substitutions of a lysine in the putative first transmembrane domain or a glutamine in the periplasmic loop caused pronounced shifts in the mechano-sensitivity curves and/or large changes in the kinetics of channel gating, suggesting that the conformational structure in these regions is critical for normal mechanosensitive channel gating.

Physical properties of sediment core AND-2A

Whole-core measurements of Wet Bulk Density (WBD), compressional (P)-wave velocity (Vp), and Magnetic Susceptibility were measured at a sampling interval of 1 or 2 centimetres (cm) throughout the AND-2A drill core for initial core characterisation and on-site correlation with seismic modeling to predict target-reflector depth. Measurements were made using a GEOTEK (Multi-Sensor-Core-Logger MSCL). Density and velocity standards were measured together with core runs of 3-6 metres (m) (and occasionally up to 18 m) throughout the entire depth range to monitor data quality. Drift of the magnetic susceptibility sensor was also monitored and corrected where necessary. These physical properties show a large range of values, reflecting the different nature of the various lithologies including extremely high velocity and density values in individual clasts, and the effects of cementation on porosity. A downcore increase in WBD and Vp occurs in the upper 200 m, however, no systematic trend exists at greater depths although large fluctuations on a m-decimetre- (dm) scale occur. Magnetic susceptibility is generally low (<100 x 10-5 SI), however, four intervals of high (>600 x 10-5 SI) susceptibility occur at 560, 800, 980 and 1 080 mbsf, indicating a relatively greater contribution of volcanic-derived material to the core site in the lower half of the AND-2A core.