Specificity of signal peptide recognition in Tat-dependent bacterial protein translocation
Specificity of signal peptide recognition in Tat-dependent bacterial protein translocation
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DOI：
10.1128/jb.183.2.604-610.2001
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发表时间：
2001-01-01
影响因子：
3.2
通讯作者：
Wiegert, T
中科院分区：
文献类型：
--
作者：
Blaudeck, N;Sprenger, GA;Wiegert, T
细菌的双胞胎精氨酸转位(TAT)途径通过细胞质膜折叠蛋白转运，在大多数情况下，这些蛋白包含一个紧密结合的辅因子。特定的氨基端信号肽具有保守的氨基酸共有基序SIT-R-R-X-F-L-K，将这些蛋白定向到TAT转运子。运动发酵单胞菌葡萄糖-果糖氧化还原酶(GFOR)是一种以NADP为辅因子的周质酶，它是以氨基末端信号肽为细胞质前体合成的，具有典型的双胞胎精氨酸信号肽的所有特征。然而，gfor在异源宿主大肠杆菌中表达时不会输出到周质，而在细胞质中发现了具有酶活性的pregfor，通过由前三甲胺N-氧化物还原酶(TMAO)还原酶(TORA)衍生的真正的E.coliTAT信号肽精确地取代pregfor信号肽，允许gfor连同其结合的辅因子输出到大肠杆菌周质，这种输出可以被羰基氰化物m-氯苯肼抑制，但不能被叠氮钠阻止，并且在E.coliatC和talae突变株中被阻止。结果表明，Tora-gfor融合蛋白的膜转位是通过TAT途径发生的，而不是通过SEC途径。此外，紧密的辅因子结合(因此正确的折叠)被发现是融合蛋白正确转位的先决条件。这些结果有力地表明，TAT信号肽并不是被不同的TAT转位酶所普遍识别的，这意味着依赖于TAT的前体蛋白的信号肽只适合于它们的同源输出装置。这种情况与已知的依赖SEC的蛋白质转位的情况形成鲜明对比。
The bacterial twin arginine translocation (Tat) pathway translocates across the cytoplasmic membrane folded proteins which, in most cases, contain a tightly bound cofactor. Specific amino-terminal signal peptides that exhibit a conserved amino acid consensus motif, SIT-R-R-X-F-L-K, direct these proteins to the Tat translocon. The glucose-fructose oxidoreductase (GFOR) of Zymomonas mobilis is a periplasmic enzyme with tightly bound NADP as a cofactor, It is synthesized as a cytoplasmic precursor with an amino-terminal signal peptide that shows all of the characteristics of a typical twin arginine signal peptide. However, GFOR is not exported to the periplasm when expressed in the heterologous host Escherichia coli, and enzymatically active pre-GFOR is found in the cytoplasm, A precise replacement of the pre-GFOR signal peptide by an authentic E. coli Tat signal peptide, which is derived from pre-trimethylamine N-oxide (TMAO) reductase (TorA), allowed export of GFOR, together with its bound cofactor, to the E. coli periplasm, This export was inhibited by carbonyl cyanide m-chlorophenylhydrazone, but not by sodium azide, and was blocked in E. coli tatC and tatAE mutant strains, Showing that membrane translocation of the TorA-GFOR fusion protein occurred via the Tat pathway and not via the Sec pathway. Furthermore, tight cofactor binding (and therefore correct folding) was found to be a prerequisite for proper translocation of the fusion protein. These results strongly suggest that Tat signal peptides are not universally recognized by different Tat translocases, implying that the signal peptides of Tat-dependent precursor proteins are optimally adapted only to their cognate export apparatus. Such a situation is in marked contrast to the situation that is known to exist for Sec-dependent protein translocation.