从Sf9细胞中纯化得到的经去污剂溶解的Patched蛋白无法与同源的Hedgehog或Ihog同源物发生强烈相互作用。
Detergent-solubilized Patched purified from Sf9 cells fails to interact strongly with cognate Hedgehog or Ihog homologs.
作者信息
Cleveland Thomas E, McCabe Jacqueline M, Leahy Daniel J
机构信息
Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, 725 N. Wolfe St., Baltimore, MD 21205, USA.
Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, 725 N. Wolfe St., Baltimore, MD 21205, USA.
出版信息
Protein Expr Purif. 2014 Dec;104:92-102. doi: 10.1016/j.pep.2014.09.012. Epub 2014 Sep 26.
Patched (Ptc) is a twelve-pass transmembrane protein that functions as a receptor for the Hedgehog (Hh) family of morphogens. In addition to Ptc, several accessory proteins including the CDO/Ihog family of co-receptors are necessary for proper Hh signaling. Structures of Hh proteins bound to members of the CDO/Ihog family are known, but the nature of the full Hh receptor complex is not well understood. We have expressed the Drosophila Patched and Mouse Patched-1 proteins in Sf9 cells and find that Sonic Hedgehog will bind to Mouse Patched-1 in isolated Sf9 cell membranes but that purified, detergent-solubilized Ptc proteins do not interact strongly with cognate Hh and CDO/Ihog homologs. These results may reflect a nonnative conformation of detergent-solubilized Ptc or that an additional factor or factors lost during purification are required for high-affinity Ptc binding to Hh.
patched（Ptc）是一种具有12个跨膜结构域的蛋白质，作为形态发生素刺猬（Hh）家族的受体发挥作用。除了Ptc之外，包括CDO/Ihog共受体家族在内的几种辅助蛋白对于正常的Hh信号传导也是必需的。已知与CDO/Ihog家族成员结合的Hh蛋白的结构，但完整的Hh受体复合物的性质尚不清楚。我们在Sf9细胞中表达了果蝇Ptc和小鼠Ptc-1蛋白，发现音猬因子（Sonic Hedgehog）将在分离的Sf9细胞膜中与小鼠Ptc-1结合，但纯化的、用去污剂溶解的Ptc蛋白与同源Hh和CDO/Ihog同源物的相互作用并不强烈。这些结果可能反映了用去污剂溶解的Ptc的非天然构象，或者是纯化过程中丢失的一种或多种额外因子是Ptc与Hh高亲和力结合所必需的。
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