Department of Molecular and Cellular Biology, Harvard University, 52 Oxford Street, Cambridge, MA 02138, USA.
Biochemistry. 2012 Aug 7;51(31):6195-206. doi: 10.1021/bi300279b. Epub 2012 Jul 25.
The TRPV4 calcium-permeable cation channel plays important physiological roles in osmosensation, mechanosensation, cell barrier formation, and bone homeostasis. Recent studies reported that mutations in TRPV4, including some in its ankyrin repeat domain (ARD), are associated with human inherited diseases, including neuropathies and skeletal dysplasias, probably because of the increased constitutive activity of the channel. TRPV4 activity is regulated by the binding of calmodulin and small molecules such as ATP to the ARD at its cytoplasmic N-terminus. We determined structures of ATP-free and -bound forms of human TRPV4-ARD and compared them with available TRPV-ARD structures. The third inter-repeat loop region (Finger 3 loop) is flexible and may act as a switch to regulate channel activity. Comparisons of TRPV-ARD structures also suggest an evolutionary link between ARD structure and ATP binding ability. Thermal stability analyses and molecular dynamics simulations suggest that ATP increases stability in TRPV-ARDs that can bind ATP. Biochemical analyses of a large panel of TRPV4-ARD mutations associated with human inherited diseases showed that some impaired thermal stability while others weakened ATP binding ability, suggesting molecular mechanisms for the diseases.
瞬时受体电位香草酸亚型 4（TRPV4）钙离子通透阳离子通道在渗透压感知、机械感觉、细胞屏障形成和骨骼动态平衡中发挥重要的生理作用。最近的研究报告称，TRPV4 的突变，包括其锚蛋白重复结构域（ARD）中的一些突变，与人类遗传性疾病有关，包括神经病变和骨骼发育不良，可能是由于通道的组成型活性增加所致。TRPV4 的活性受到细胞质 N 端 ARD 上钙调蛋白和小分子（如 ATP）与 ARD 结合的调节。我们测定了无 ATP 和有 ATP 结合形式的人 TRPV4-ARD 结构，并将其与现有的 TRPV-ARD 结构进行了比较。第三内环重复区（Finger 3 loop）具有柔性，可能作为调节通道活性的开关。TRPV-ARD 结构的比较还表明，ARD 结构与 ATP 结合能力之间存在进化联系。热稳定性分析和分子动力学模拟表明，ATP 增加了能够结合 ATP 的 TRPV-ARD 的稳定性。对与人类遗传性疾病相关的大量 TRPV4-ARD 突变的生化分析表明，一些突变会降低热稳定性，而另一些则削弱了 ATP 结合能力，这提示了这些疾病的分子机制。
