Interhelical hydrogen bonds in the CFTR membrane domain.
Critical mutations in the membrane-spanning domains of proteins cause many human diseases. We report the expression in Escherichia coli of helix-loop-helix segments of the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel domain in milligram quantities. Analysis of gel migration patterns of these constructs, in conjunction with circular dichroism spectroscopy, demonstrate that a neutral-to-charged, CF-phenotypic point mutation of a hydrophobic residue (V232D) in the CFTR transmembrane (TM) helix 4 induces a hydrogen bond with neighboring wild type Gln 207 in TM helix 3. As an electrostatic crosslink within a hydrocarbon phase, such a hydrogen bond could alter the normal assembly and alignment of CFTR TM helices and/or impede their movement in response to substrate transport. Our results imply that membrane proteins may be vulnerable to loss of function through formation of membrane-buried interhelical hydrogen bonds by partnering of proximal polar side chains.
Therien
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Division of Structural Biology and Biochemistry, Research Institute, Hospital for Sick Children, 555 University Avenue, Toronto, Ontario M5G 1X8, Canada.
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Deber
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D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D002942:Circular Dichroism; D003198:Computer Simulation; D003550:Cystic Fibrosis / Q000235:genetics; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D003902:Detergents / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D018257:Helix-Loop-Helix Motifs; D006801:Humans; D006860:Hydrogen Bonding; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010641:Phenotype; D017354:Point Mutation / Q000235:genetics; D017433:Protein Structure, Secondary / Q000187:drug effects; D017434:Protein Structure, Tertiary; D055672:Static Electricity; D013816:Thermodynamics
C505032:CFTR protein, human; D003902:Detergents; D019005:Cystic Fibrosis Transmembrane Conductance Regulator