title: Molecular determinants of permeation through the cation channel TRPM6.
abstract: TRPM6 and its closest relative TRPM7 are members of the Transient Receptor Potential Melastatin (TRPM) subfamily of cation channels and are known to be Mg2+ permeable. By aligning the sequence of the putative TRPM6 pore with the pore sequences of the other subfamily members, we located in the loop between the fifth and the sixth transmembrane domain, a stretch of amino acids residues, 1028GEIDVC1033, as the potential selectivity filter. Two negatively charged residues, E1024 (conserved in TRPM6, TRPM7, TRPM1 and TRPM3) and D1031 (conserved along the entire TRPM subfamily), were identified as important determinants of cation permeation through TRPM6, because neutralization of both residues into an alanine resulted in non-functional channels. Neutralization of E1029 (conserved in TRPM6, TRPM7, TRPM4 and TRPM5) resulted in channels with increased conductance for Ba2+ and Zn2+, decreased ruthenium red sensitivity and larger pore diameter compared to wild-type TRPM6. Changing the residue I1030 into methionine, resulted in channels with lower conductance for Ni2+, decreased sensitivity to ruthenium red block and reduced pore diameter. Thus, these data demonstrate that amino acid residues E1024, I1030 and D1031 are important for channel function and that subtle amino acid variation in the pore region accounts for TRPM6 permeation properties.
authors.affiliation: Department of Physiology, Nijmegen Centre for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, The Netherlands.
authors.forename: Catalin N
authors.initials: CN
authors.lastname: Topala
authors.forename: Wouter Tiel
authors.initials: WT
authors.lastname: Groenestege
authors.forename: Stéphanie
authors.initials: S
authors.lastname: Thébault
authors.forename: Dennis
authors.initials: D
authors.lastname: van den Berg
authors.forename: Bernd
authors.initials: B
authors.lastname: Nilius
authors.forename: Joost G
authors.initials: JG
authors.lastname: Hoenderop
authors.forename: René J
authors.initials: RJ
authors.lastname: Bindels
mesh_terms: D000595:Amino Acid Sequence; D002412:Cations / Q000378:metabolism; D002460:Cell Line; D006801:Humans; D017136:Ion Transport; D008274:Magnesium / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D018408:Patch-Clamp Techniques; D012430:Ruthenium Red / Q000494:pharmacology; D016415:Sequence Alignment; D012964:Sodium / Q000378:metabolism; D050053:TRPM Cation Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
chemical_list: D002412:Cations; D050053:TRPM Cation Channels; C459082:TRPM6 protein, human; D012430:Ruthenium Red; D012964:Sodium; D008274:Magnesium