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Biophys Chem. 2010 May;148(1-3):112-20. doi: 10.1016/j.bpc.2010.03.004. Epub 2010 Mar 16.

In vitro association of fragments of a beta-sheet membrane protein.

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Department of Molecular Biology, University of Aarhus, Gustav Wieds Vej 10C, DK-8000 Aarhus C, Denmark.


Although the beta-barrel membrane protein OmpA can be produced in a biologically active form in E. coli from co-expressed fragments, the fragments have not been demonstrated to associate in vitro. We have produced 3 complementary fragment pairs of OmpA which can associate to form a folded complex according to the SDS band-shift assay. We are able to convert 25-35% of the fragment populations to non-covalent but SDS-stable complexes. The periplasmic chaperone Skp effectively prevented this association. Two separately expressed and purified overlapping fragments of OmpA can form a protease-resistant complex that undergoes the characteristic band-shift upon heating. Our work demonstrates that although membrane insertion and folding of beta-barrel membrane proteins may be a cooperative process, the fragments can associate in vitro without any additional components. However, the low yield and slow folding rates indicate that partially unfolded or destabilized beta-sheet membrane proteins can potentially engage in many non-native interactions.

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