Tinker, Tailor

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Science  01 Oct 2004:
Vol. 306, Issue 5693, pp. 21
DOI: 10.1126/science.306.5693.21c

The rarity of naturally enriched sources of eukaryotic membrane proteins has made it difficult to pursue classical biochemistry—purification of detergent-solubilized transporters and receptors—to the end point of crystallization and structure determination. Membrane proteins are happiest within their native lipid environment, which, given the diversity of lipid molecules, is antithetical to the homogeneity of a crystal. However, a series of studies on the well-characterized bacterial mechanosensitive channel MscL tackles these challenges and offers some hope for future applications.

Clayton et al. divided the 15-kD protein into thirds and synthesized the three peptide segments chemically before ligating them into a full-length molecule in the absence of detergents and lipids. Reconstitution into vesicles and patch-clamp measurements confirmed that pressure-sensitive channels had in fact been made. Berrier et al. used a coupled in vitro transcription-translation system to make milligram quantities of MscL in the absence of membranes and were also able to demonstrate activity of the product. Finally, Becker et al. have converted MscL into a completely soluble form by attaching polyethylene- glycol-polyamide tails at four places where the protein would face the hydrophobic interior of the lipid bilayer. Although activity can no longer be measured directly, the structure is by several measures the same as that of the natural protein. — GJC

Proc. Natl. Acad. Sci. U.S.A. 101, 4764 (2004); Biochemistry 10.1021/bi049049y (2004); J. Mol. Biol. 10.1016/j.jmb.2004.08.062 (2004).

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