SecA functions in vivo as a discrete anti-parallel dimer to promote protein transport.
SecA functions in vivo as a discrete anti-parallel dimer to promote protein transport.
复制标题
SECA在体内发挥离散的抗平行二聚体，可促进蛋白质转运。
DOI：
10.1111/mmi.13567
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发表时间：
2017-02
影响因子：
3.6
通讯作者：
Oliver D
中科院分区：
文献类型：
--
作者：
Banerjee T;Lindenthal C;Oliver D
SecA ATP 酶马达蛋白通过结合底物蛋白和 SecY 通道复合物并利用其 ATP 酶活性驱动蛋白质跨质膜易位，在细菌蛋白质转运中发挥核心作用。 SecA 已被证明存在于易位配体调节的动态单体-二聚体平衡中，并且二聚体的多种结构形式已结晶。由于二聚体的结构形式仍然是一个有争议且未解决的问题，我们通过沿着对应于五种不同 SecA X 射线结构的二聚体界面设计 ρ-苯甲酰基苯丙氨酸并评估它们的体内光交联模式来解决这个问题。无论 SecA 是胞质还是脂质或 SecYEG 结合状态，离散的反平行 1M6N 样二聚体是体内发现的主要二聚体（如果不是唯一的二聚体）。与稳定易位中间体结合的 SecA 在体内与第二个 SecA 原聚体在其 1M6N 界面交联，表明这种特定的二聚体可能促进活性蛋白易位。总而言之，我们的研究强化了至少部分假设 SecA 二聚体驱动的易位机制的模型。 SecA ATPase 促进蛋白质通过整合膜 SecY 通道复合体转运。通过将位点特异性交联剂设计到潜在的二聚体界面并进行体内光交联，探索了已结晶的各种 SecA 二聚体的生理形式。结果表明，细胞内存在单个离散的二聚体物质，并且该物质也在利用 SecA-OmpA-GFP 三聚体蛋白通过 SecY 通道的阻滞蛋白转运过程中被捕获。
SecA ATPase motor protein plays a central role in bacterial protein transport by binding substrate proteins and the SecY channel complex and utilizing its ATPase activity to drive protein translocation across the plasma membrane. SecA has been shown to exist in a dynamic monomer-dimer equilibrium modulated by translocation ligands, and multiple structural forms of the dimer have been crystallized. Since the structural form of the dimer remains a controversial and unresolved question, we addressed this matter by engineering ρ-benzoylphenylalanine along dimer interfaces corresponding to the five different SecA x-ray structures and assessing their in vivo photo-crosslinking pattern. A discrete anti-parallel 1M6N-like dimer was the dominant if not exclusive dimer found in vivo, whether SecA was cytosolic or in lipid or SecYEG-bound states. SecA bound to a stable translocation intermediate was crosslinked in vivo to a second SecA protomer at its 1M6N interface, suggesting that this specific dimer likely promotes active protein translocation. Taken together, our studies strengthen models that posit, at least in part, a SecA dimer-driven translocation mechanism. SecA ATPase facilitates protein transport through the integral membrane SecY channel complex. The physiological form of the various SecA dimers that have been crystallized was explored by engineering a site-specific crosslinker into potential dimer interfaces and performing in vivo photo-crosslinking. The results indicate that a single discrete dimer species is present within the cell, and this species was also captured during arrested protein transport through the SecY channel utilizing a SecA-OmpA-GFP trimeric protein.