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  • Title: Cystic fibrosis transmembrane conductance regulator: expression and helicity of a double membrane-spanning segment.
    Author: Peng S, Liu LP, Emili AQ, Deber CM.
    Journal: FEBS Lett; 1998 Jul 10; 431(1):29-33. PubMed ID: 9684859.
    Abstract:
    The gene responsible for cystic fibrosis encodes a membrane protein--the 1480-residue cystic fibrosis transmembrane conductance regulator (CFTR)--in which membrane-based CF-phenotypic mutants alter pore structure and/or impair ion transport. We report the preparation in milligram quantities and conformational characterization of a polypeptide comprised of CFTR transmembrane (TM) segments 3-4, a putative 'helical hairpin' portion of the CFTR TM1-6 domain. The TM segment 3-4 of CFTR was expressed in E. coli as a fusion protein linked to the C-terminus of His-tagged thioredoxin. Nickel chelate affinity chromatography, followed by release from the carrier by digestion with thrombin protease, gave free CFTR(TM3-4). Monitoring of the folding properties and conformational state(s) of the TM3-4 polypeptide using circular dichroism spectroscopy indicated a partial alpha-helical conformation in aqueous buffer, with up to 30% increase in alpha-helical content observed in membrane-mimetic environments.
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