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

D Debnath, K L Nielsen, D E Otzen

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaperJournal articleResearchpeer-review


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.
Original languageEnglish
Book seriesAdvances in Biophysical Chemistry
Pages (from-to)112-20
Number of pages9
Publication statusPublished - 1 May 2010


  • Amino Acid Sequence
  • Bacterial Outer Membrane Proteins
  • DNA-Binding Proteins
  • Escherichia coli Proteins
  • Hot Temperature
  • Kinetics
  • Models, Molecular
  • Molecular Chaperones
  • Peptide Hydrolases
  • Protein Folding
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Substrate Specificity


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