Identification and Tetramer Structure of Hemin-Binding Protein SPD_0310 Linked to Iron Homeostasis and Virulence of Streptococcus pneumoniae.
Identification and Tetramer Structure of Hemin-Binding Protein SPD_0310 Linked to Iron Homeostasis and Virulence of Streptococcus pneumoniae.
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DOI：
10.1128/msystems.00221-22
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发表时间：
2022-06-28
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影响因子：
6.4
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中科院分区：
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铁和含铁化合物是细菌毒力和宿主感染所必需的。氯化血红素是一种重要的补充化合物，在缺铁的环境中细菌的生存。尽管对氯化血红素的代谢有着浓厚的兴趣，但氯化血红素在革兰氏阳性菌中转运的详细机制尚未见报道。结果表明，SPD_0310的同源蛋白在革兰氏阳性菌中具有显著的保守性（P < 0.001），属于一个未知蛋白家族（UPF 0371）。热力学和动力学研究的结果表明，SPD_0310具有高的氯化血红素结合亲和力。有趣的是，我们发现SPD_0310的晶体结构呈现同源四聚体构象，这是氯化血红素结合所必需的。SPD_0310可以与位于细胞表面的许多氯化血红素结合蛋白（SPD_0090、SPD_1609和GAPDH）相互作用，这有助于氯化血红素转移到细胞质中。它还与细胞质中的其他铁转运蛋白（SPD_0226和SPD_0227）具有高亲和力，这有助于细胞中的铁再分布。更重要的是，spd_0310基因的敲除（Δspd_0310）导致铁含量和许多细菌粘附因子的蛋白表达水平降低。此外，动物模型显示Δspd_0310菌株具有比野生型更低的毒力。基于晶体学和生化研究，我们推测SPD_0310是一种氯化血红素中间转运体，有助于铁稳态，并进一步影响肺炎链球菌在宿主中的毒力。我们的研究不仅为深入阐明氯化血红素在细菌中的转运机制提供了重要的理论基础，而且为开发基于金属转运系统的新型抗菌剂提供了重要的候选靶点。重要性铁是细菌毒力和宿主感染的必需元素。革兰氏阳性菌中氯化血红素代谢的详细研究很少。SPD_0310属于UPF 0371蛋白家族，同源性分析和进化树分析结果表明，SPD_0310在革兰氏阳性菌中分布广泛且高度保守。然而，UPF 0371家族的功能仍然未知。我们成功地确定了apo-SPD_0310的晶体结构，它是一种同源四聚体。我们发现胞质蛋白SPD_0310具有特殊的四聚体结构，具有很强的氯化血红素结合能力，并与许多铁转运蛋白相互作用，从而促进氯化血红素从细胞外空间转移到细胞质。详细的功能分析结果表明，SPD_0310在动物体内可能作为类似于血红蛋白的氯化血红素转运蛋白发挥作用，并有助于细菌铁稳态和毒力。该研究为开发抗致病性革兰氏阳性菌的抗菌药物提供了新的靶点。
Iron and iron-containing compounds are essential for bacterial virulence and host infection. Hemin is an important supplement compound for bacterial survival in an iron-deficient environment. Despite strong interest in hemin metabolism, the detailed mechanism of hemin transportation in Gram-positive bacteria is yet to be reported. The results of our study revealed that the homologous proteins of SPD_0310 were significantly conservative in Gram-positive bacteria (P < 0.001), and these proteins were identified as belonging to an uncharacterized protein family (UPF0371). The results of thermodynamic and kinetic studies have shown that SPD_0310 has a high hemin-binding affinity. Interestingly, we found that the crystal structure of SPD_0310 presented a homotetramer conformation, which is required for hemin binding. SPD_0310 can interact with many hemin-binding proteins (SPD_0090, SPD_1609, and GAPDH) located on the cell surface, which contributes to hemin transfer to the cytoplasm. It also has a high affinity with other iron transporters in the cytoplasm (SPD_0226 and SPD_0227), which facilitates iron redistribution in cells. More importantly, the knockout of the spd_0310 gene (Δspd_0310) resulted in a decrease in the iron content and protein expression levels of many bacterial adhesion factors. Moreover, the animal model showed that the Δspd_0310 strain has a lower virulence than the wild type. Based on the crystallographic and biochemical studies, we inferred that SPD_0310 is a hemin intermediate transporter which contributes to iron homeostasis and further affects the virulence of Streptococcus pneumoniae in the host. Our study provides not only an important theoretical basis for the in-depth elucidation of the hemin transport mechanism in bacteria but also an important candidate target for the development of novel antimicrobial agents based on metal transport systems. IMPORTANCE Iron is an essential element for bacterial virulence and infection of the host. The detailed hemin metabolism in Gram-positive bacteria has rarely been studied. SPD_0310 belongs to the UPF0371 family of proteins, and results of homology analysis and evolutionary tree analysis suggested that it was widely distributed and highly conserved in Gram-positive bacteria. However, the function of the UPF0371 family remains unknown. We successfully determined the crystal structure of apo-SPD_0310, which is a homotetramer. We found that cytoplasmic protein SPD_0310 with a special tetramer structure has a strong hemin-binding ability and interacts with many iron transporters, which facilitates hemin transfer from the extracellular space to the cytoplasm. The results of detailed functional analyses indicated that SPD_0310 may function as a hemin transporter similar to hemoglobin in animals and contributes to bacterial iron homeostasis and virulence. This study provides a novel target for the development of antimicrobial drugs against pathogenic Gram-positive bacteria.
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影响因子：
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作者：
Liu X;Gong J;Wei T;Wang Z;Du Q;Zhu D;Huang Y;Xu S;Gu L
通讯作者：
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影响因子：
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作者：
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DOI：
10.1107/s0907444904019158
发表时间：
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影响因子：
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