Sec-dependent membrane protein biogenesis: SecYEG, preprotein hydrophobicity and translocation kinetics control the stop-transfer function
Sec-dependent membrane protein biogenesis: SecYEG, preprotein hydrophobicity and translocation kinetics control the stop-transfer function
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DOI：
10.1093/emboj/17.3.696
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发表时间：
1998-02-02
期刊：
影响因子：
11.4
通讯作者：
Wickner, W
中科院分区：
文献类型：
--
作者：
Duong, F;Wickner, W
蛋白前转位酶催化膜蛋白整合和完全易位。膜蛋白必须中断其易位，并从转位酶侧释放到脂质双分子层。我们通过体外反应分析了大肠杆菌前蛋白转位酶的易位阻滞和侧向释放活性，前蛋白proOmpA携带合成的停止转移序列，膜蛋白整合是催化性的，其发生动力学与proOmpA本身相似，只需要SecYEG和SecA的功能。虽然强疏水片段会引导蛋白质离开转位酶进入脂质双分子层，但具有中间疏水片段的蛋白质在易位状态和膜整合状态之间平均分配。PMF、不同ATP浓度或合成易位阻滞的影响分析表明，轻度疏水片段的停止易位效率取决于易位动力学。相反，从转位酶到脂质的侧向分配完全取决于温度，不需要SecA ATP水解或SecA膜循环，因此易位阻滞由SecYEG转位酶活性控制，而侧向释放和膜整合由片段本身的疏水性指导。我们的研究结果表明，有效的易位阻滞需要比侧向释放到膜上更大的疏水性。
Preprotein translocase catalyzes membrane protein integration as well as complete translocation. Membrane proteins must interrupt their translocation and be laterally released from the translocase into the lipid bilayer. We have analyzed the translocation arrest and lateral release activities of Escherichia coli preprotein translocase with an in vitro reaction and the preprotein proOmpA carrying a synthetic stop-transfer sequence, Membrane protein integration is catalytic, occurs with kinetics similar to those of proOmpA itself and only requires the functions of SecYEG and SecA. Though a strongly hydrophobic segment will direct the protein to leave the translocase and enter the lipid bilayer, a protein with a segment of intermediate hydrophobicity partitions equally between the translocated and membrane-integrated states, Analysis of the effects of PMF, varied ATP concentrations or synthetic translocation arrest show that the stop-translocation efficiency of a mildly hydrophobic segment depends on the translocation kinetics. In contrast, the lateral partitioning from translocase to lipids depends solely on temperature and does not require SecA ATP hydrolysis or SecA membrane cycling, Thus translocation arrest is controlled by the SecYEG translocase activity while lateral release and membrane integration are directed by the hydrophobicity of the segment itself, Our results suggest that a greater hydrophobicity is required for efficient translocation arrest than for lateral release into the membrane.