We found the activities and later cloned the gene of the Mechanosensitive channel of Large conductance, MscL, of E. coli. Although mechanosensitive (MS) channels are found in many systems and play roles in the senses of touch, hearing, etc., MscL and MscS (below) are the few tangible models to study the biochemical and biophysical details of mechanosensation.
We first encountered MS channel activities in a patch-clamp survey of the surface of giant cells generated from bacteria (1) (Fig. 1). These activities could be biochemically separated by gel filtration (2). Using reconstitution as an assay (patch clamping reconstituted fractions), the MscL protein was identified, and the corresponding gene cloned (3).
Gain-of-function mutants with "loose-cannon" channel activities (4), site-directed mutations (5,6), and biochemical studies (7) led to a model of the MscL-channel subunit having two-transmembrane alpha helices (TM1 and TM2) (8). The Rees laboratory (9) solved a crystal structure of the MscL homolog in Mycobacteium tuberculosis. The channel is a homopentamer, with the five TM1's coming together towards the cytoplasm to close the pore (Fig. 2). The open structure has been deduced by a variety of studies by other laboratories.
The Booth laboratory cloned the second MS-channel gene (mscS), (10), and showed that the mscL- mscS- double mutant bacteria lyse upon an osmotic downshock (Fig. 3). This work supports the previous hypothesis that these MS channels function as emergency valves. Upon downshocks, they open to jettison solutes, reducing the turgor and thereby preventing the bacteria from bursting (Fig. 3).
See (11, 12, 13) for reviews.
1. Martinac et al. (1987 ) PNAS 84: 2297.
2. Sukharev et al. 1993) Biophy. J. 65: 1.
3. Sukharev et al. (1994) Nature 368: 265
4. Ou et al. (1998) PNAS 95: 11471.
5. Yoshimura et al. (1999) Biophy. J. 77: 144.
6. Yoshimura et al. (2001) Biophy. J. 80: 2198.
7. Blount et al. (1996) EMBO J. 15: 4798.
8. Sukharev et al. (1997) Annu. Rev. Physiol. 59: 633.
9. Chang et al. (1998) Science 282: 2220.
10. Levina et al. (1998) EMBO J. 18: 1730.
11. Sukharev and Corey (2004) Science STKE. re 4.
12. Anishkin and Kung (2005) Cur. Opin. Neurobiol. 15: 397.
13. Kung (2005) Nature 436: 647.