Schreiner Madeleine, Niemann Hartmut H
Department of Chemistry, Bielefeld University, PO Box 10 01 31, 33501 Bielefeld, Germany.
BMC Struct Biol. 2012 Jun 18;12:13. doi: 10.1186/1472-6807-12-13.
Type III secretion systems are used by Gram-negative bacteria as "macromolecular syringes" to inject effector proteins into eukaryotic cells. Two hydrophobic proteins called translocators form the necessary pore in the host cell membrane. Both translocators depend on binding to a single chaperone in the bacterial cytoplasm to ensure their stability and efficient transport through the secretion needle. It was suggested that the conserved chaperones bind the more divergent translocators via a hexapeptide motif that is found in both translocators and conserved between species.
We crystallized a synthetic decapeptide from the Yersinia enterocolitica minor type III secretion translocator YopD bound to its cognate chaperone SycD and determined the complex structure at 2.5 Å resolution. The structure of peptide-bound SycD is almost identical to that of apo SycD with an all helical fold consisting of three tetratricopeptide repeats (TPRs) and an additional C-terminal helix. Peptide-bound SycD formed a kinked head-to-head dimer that had previously been observed for the apo form of SycD. The homodimer interface comprises both helices of the first tetratricopeptide repeat. The YopD peptide bound in extended conformation into a mainly hydrophobic groove on the concave side of SycD. TPRs 1 and 2 of SycD form three hydrophobic pockets that accommodated the conserved hydrophobic residues at position 1, 3 and 6 of the translocator hexapeptide sequence. Two tyrosines that are highly conserved among translocator chaperones contribute to the hydrophobic patches but also form hydrogen bonds to the peptide backbone.
The interaction between SycD and YopD is very similar to the binding of the Pseudomonas minor translocator PopD to its chaperone PcrH and the Shigella major translocator IpaB to its chaperone IpgC. This confirms the prediction made by Kolbe and co-workers that a hexapeptide with hydrophobic residues at three positions is a conserved chaperone binding motif. Because the hydrophobic groove on the concave side of translocator chaperones is involved in binding of the major and the minor translocator, simultaneous binding of both translocators to a single type III secretion class II chaperone appears unlikely.
革兰氏阴性菌利用III型分泌系统作为“大分子注射器”，将效应蛋白注入真核细胞。两种称为转运体的疏水蛋白在宿主细胞膜上形成必要的孔道。这两种转运体都依赖于与细菌细胞质中的单个伴侣蛋白结合，以确保其稳定性并通过分泌针高效运输。有人提出，保守的伴侣蛋白通过六肽基序与差异较大的转运体结合，该六肽基序在两种转运体中均有发现且在物种间保守。
我们使来自小肠结肠炎耶尔森菌次要III型分泌转运体YopD的合成十肽与其同源伴侣蛋白SycD结合并结晶，以2.5 Å的分辨率确定了复合物结构。肽结合的SycD结构几乎与游离SycD相同，具有由三个四肽重复序列（TPR）和一个额外的C末端螺旋组成的全螺旋折叠。肽结合的SycD形成了一个扭结的头对头二聚体，之前在游离形式的SycD中也观察到过。同型二聚体界面包含第一个四肽重复序列的两个螺旋。YopD肽以伸展构象结合到SycD凹面的一个主要疏水凹槽中。SycD的TPR 1和TPR 2形成三个疏水口袋，容纳转运体六肽序列第1、3和6位的保守疏水残基。在转运体伴侣蛋白中高度保守的两个酪氨酸有助于形成疏水区域，但也与肽主链形成氢键。
SycD与YopD之间的相互作用与绿脓杆菌次要转运体PopD与其伴侣蛋白PcrH以及痢疾志贺菌主要转运体IpaB与其伴侣蛋白IpgC的结合非常相似。这证实了科尔贝及其同事的预测，即三个位置带有疏水残基的六肽是保守的伴侣蛋白结合基序。由于转运体伴侣蛋白凹面的疏水凹槽参与主要和次要转运体的结合，因此两种转运体同时结合到单个III型分泌II类伴侣蛋白上似乎不太可能。
