Factors involved in the stop transfer process in membrane protein biogenesis

(1)我们构建了一个膜蛋白模型proOmpAl65-Lacy(TM12)-PhoA，并分析了SecY在其整合到膜中的作用。我们发现，由于在secY125突变株中进行了低效的strop转移反应，该模型蛋白不太稳定地整合到膜中。而在PriA(Secy)4-1突变株中，随着效率的提高，发生了停止转移反应。这些结果有力地表明，SecY直接参与了膜蛋白生物发生中的停止转移过程。(第二年)我们以前曾提出FtsH参与停止转移过程。我们发现，1)FtsH与变性碱性磷酸酶有亲和力，2)FtsH与变性碱性磷酸酶的结合不依赖于ATP，3)FtsH不能降解结合的碱性磷酸酶。FtsH的这种变性蛋白结合活性可能在膜蛋白的生物发生，特别是停止转移过程中发挥一定的作用。

(1st year) We constructed a model membrane protein, proOmpAl65-LacY(TM12)-PhoA, and analyzed the roles of SecY in its integration into the membrane. We found that this model protein was less stably integrated into the membrane as a result of inefficient strop transfer reaction in the secY125 mutant strain than in the wild type strain. On the other hand, the stop transfer reaction occurred with the enhanced efficiency in the prIA(secY) 4-1 mutant strain. These results strongly suggest that SecY is directly involved in the stop transfer process in biogenesis of membrane proteins.(2nd year) We suggested previously that FtsH is involved in the stop transfer process. We found that, 1) FtsH has affinity to a denatured alkaline phosphatase, 2) the binding of FtsH to a denatured alkaline phosphatase is independent of ATP, 3) FtsH is unable to degrade bound alkaline phosphatase. This denatured protein binding activity of FtsH may have some role in the biogenesis of membrane proteins, especially the stop transfer process.