A multi-pronged search for a common structural motif in the secretion signal of Salmonella enterica serovar Typhimurium type III effector proteins.
A multi-pronged search for a common structural motif in the secretion signal of Salmonella enterica serovar Typhimurium type III effector proteins.
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DOI：
10.1039/c0mb00097c
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发表时间：
2010-12
影响因子：
--
通讯作者：
McDermott JE
中科院分区：
文献类型：
--
作者：
Buchko GW;Niemann G;Baker ES;Belov ME;Smith RD;Heffron F;Adkins JN;McDermott JE
许多致病性革兰氏阴性细菌使用III型分泌系统（T3 SS）将效应蛋白递送到宿主细胞中，在那里它们重新编程宿主防御并促进发病。前20-30个N-末端残基通常含有靶向效应蛋白易位的“分泌信号”，然而，从未发现共有序列基序。最近的机器学习方法，如基于支持向量机（SVM）的毒力效应子鉴定和评价（SIEVE），已经提高了从基因组序列信息中鉴定效应子蛋白的能力。虽然这些方法都表明T3 SS分泌信号具有特征性氨基酸组成偏差，但仍不清楚氨基酸模式是否重要以及是否存在指导识别的任何统一结构特性。为了解决这些问题，合成了对应于肠道沙门氏菌血清型鼠伤寒沙门氏菌效应子SseJ的分泌信号的肽（残基1-30，SseJ），沿着产生高（SseJ-H）和低（SseJ-L）SIEVE评分的相同氨基酸组成的乱序肽。这三种肽的分泌特性进行了测试，使用分泌信号CyaA融合试验和它们的结构特性探测使用圆二色性，核磁共振，离子迁移谱-质谱。SIEVE的分泌预测与J774巨噬细胞的信号-CyaA融合实验结果相匹配，表明SseJ分泌信号具有一定的序列顺序依赖性。结构研究表明，SseJ，SseJ-H，和SseJ-L肽在水溶液中是内在无序的，只有在结构稳定剂如1，1，1，3，3，3-六氟异丙醇的存在下，才具有采用新生螺旋结构的小倾向。内源性紊乱可能是效应分泌信号的普遍特征，因为在对对应于S.鼠伤寒杆菌效应子SptP、SopD-2、GtgE和鼠疫耶尔森氏菌效应子YopH。
Many pathogenic Gram-negative bacteria use a type III secretion system (T3SS) to deliver effector proteins into the host cell where they reprogram host defenses and facilitate pathogenesis. The first 20–30 N-terminal residues usually contain the ‘secretion signal’ that targets effector proteins for translocation, however, a consensus sequence motif has never been discerned. Recent machine-learning approaches, such as support vector machine (SVM)-based Identification and Evaluation of Virulence Effectors (SIEVE), have improved the ability to identify effector proteins from genomics sequence information. While these methods all suggest that the T3SS secretion signal has a characteristic amino acid composition bias, it is still unclear if the amino acid pattern is important and if there are any unifying structural properties that direct recognition. To address these issues a peptide corresponding to the secretion signal for Salmonella enterica serovar Typhimurium effector SseJ was synthesized (residues 1–30, SseJ) along with scrambled peptides of the same amino acid composition that produced high (SseJ-H) and low (SseJ-L) SIEVE scores. The secretion properties of these three peptides were tested using a secretion signal–CyaA fusion assay and their structural properties probed using circular dichroism, nuclear magnetic resonance, and ion mobility spectrometry–mass spectrometry. The secretion predictions from SIEVE matched signal–CyaA fusion experimental results with J774 macrophages suggesting that the SseJ secretion signal has some sequence order dependence. The structural studies showed that the SseJ, SseJ-H, and SseJ-L peptides were intrinsically disordered in aqueous solution with a small predisposition to adopt nascent helical structure only in the presence of structure stabilizing agents such as 1,1,1,3,3,3-hexafluoroisopropanol. Intrinsic disorder may be a universal feature of effector secretion signals as similar conclusions were reached following structural characterization of peptides corresponding to the N-terminal regions of the S. Typhimurium effectors SptP, SopD-2, GtgE, and the Yersinia pestis effector YopH.
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