Topala Catalin N, Groenestege Wouter Tiel, Thébault Stéphanie, van den Berg Dennis, Nilius Bernd, Hoenderop Joost G, Bindels René J
Department of Physiology, Nijmegen Centre for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, The Netherlands.
Cell Calcium. 2007 Jun;41(6):513-23. doi: 10.1016/j.ceca.2006.10.003. Epub 2006 Nov 13.
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.
瞬时受体电位褪黑素（TRPM）阳离子通道亚家族的成员TRPM6及其关系最密切的TRPM7已知可通透Mg2+。通过将假定的TRPM6孔道序列与其他亚家族成员的孔道序列进行比对，我们在第五和第六个跨膜结构域之间的环中定位了一段氨基酸残基1028GEIDVC1033，作为潜在的选择性过滤器。两个带负电荷的残基E1024（在TRPM6、TRPM7、TRPM1和TRPM3中保守）和D1031（在整个TRPM亚家族中保守）被确定为阳离子通过TRPM6渗透的重要决定因素，因为将这两个残基都突变为丙氨酸会导致通道失去功能。将E1029（在TRPM6、TRPM7、TRPM4和TRPM5中保守）突变为丙氨酸后，与野生型TRPM6相比，通道对Ba2+和Zn2+的电导增加，对钌红的敏感性降低，孔径增大。将残基I1030变为甲硫氨酸后，通道对Ni2+的电导降低，对钌红阻断的敏感性降低，孔径减小。因此，这些数据表明氨基酸残基E1024、I1030和D1031对通道功能很重要，并且孔道区域中细微的氨基酸变化决定了TRPM6的渗透特性。
