Structural, Bioinformatic, and In Vivo Analyses of Two Treponema pallidum Lipoproteins Reveal a Unique TRAP Transporter
Structural, Bioinformatic, and In Vivo Analyses of Two Treponema pallidum Lipoproteins Reveal a Unique TRAP Transporter
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DOI：
10.1016/j.jmb.2012.01.015
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发表时间：
2012-03-09
影响因子：
5.6
通讯作者：
Norgard, Michael V.
中科院分区：
文献类型：
--
作者：
Deka, Ranjit K.;Brautigam, Chad A.;Norgard, Michael V.
梅毒的细菌病原体梅毒螺旋体预计会编码一种三联 ATP 独立周质转运蛋白 (TRAP-T)。 TRAP-T 通常采用周质底物结合蛋白 (SBP) 将同源配体递送至跨膜同向转运蛋白。在此，我们证明编码来自苍白球的假定 TRAP-T 组件的基因 tp0957（SBP）和 tp0958（同向转运蛋白）位于具有未表征的第三个基因 tp0956 的操纵子中。我们确定了重组Tp0956的晶体结构；该蛋白质是三聚体并有一个孔。 Tp0956 的一部分形成类似于“四肽重复”(TPR) 基序的组件。还测定了重组Tp0957的晶体结构；与其他 TRAP-T 的 SBP 一样，有两个由裂缝隔开的叶。在这些其他 SBP 中，裂口结合带负电荷的配体。然而，Tp0957 的裂口具有惊人的疏水性化学成分，表明其配体可能本质上不同并且可能是疏水性的。对 Tp0956 和 Tp0957 重组版本的分析超速离心证实这些蛋白质强烈结合。使用体内交联实验证实了天然分子的这种前所未有的相互作用。最后，生物信息学分析表明，该转运蛋白代表了 TPAT（TPR 蛋白相关 TRAP-T）的一个新亚家族，其需要包含 TPR 的辅助蛋白的作用来进行潜在疏水性配体的周质转运。 (C) 2012 Elsevier Ltd. 保留所有权利。
Treponema pallidum, the bacterial agent of syphilis, is predicted to encode one tripartite ATP-independent periplasmic transporter (TRAP-T). TRAP-Ts typically employ a periplasmic substrate-binding protein (SBP) to deliver the cognate ligand to the transmembrane symporter. Herein, we demonstrate that the genes encoding the putative TRAP-T components from T. pallidum, tp0957 (the SBP), and tp0958 (the symporter), are in an operon with an uncharacterized third gene, tp0956. We determined the crystal structure of recombinant Tp0956; the protein is trimeric and perforated by a pore. Part of Tp0956 forms an assembly similar to those of "tetratricopeptide repeat" (TPR) motifs. The crystal structure of recombinant Tp0957 was also determined; like the SBPs of other TRAP-Ts, there are two lobes separated by a cleft. In these other SBPs, the cleft binds a negatively charged ligand. However, the cleft of Tp0957 has a strikingly hydrophobic chemical composition, indicating that its ligand may be substantially different and likely hydrophobic. Analytical ultracentrifugation of the recombinant versions of Tp0956 and Tp0957 established that these proteins associate avidly. This unprecedented interaction was confirmed for the native molecules using in vivo cross-linking experiments. Finally, bioinforrnatic analyses suggested that this transporter exemplifies a new subfamily of TPATs (TPR-protein-associated TRAP-Ts) that require the action of a TPR-containing accessory protein for the periplasmic transport of a potentially hydrophobic ligand(s). (C) 2012 Elsevier Ltd. All rights reserved.