The major human pathogen encounters the immune-derived oxidant hypothiocyanous acid (HOSCN) at sites of colonization and infection. We recently identified the pneumococcal hypothiocyanous acid reductase (Har), a member of the flavoprotein disulfide reductase enzyme family, and showed that it contributes to the HOSCN tolerance of . Here, we demonstrate in mouse models of pneumococcal infection that Har is critical for colonization and invasion. In a colonization model, bacterial load was attenuated dramatically in the nasopharynx when was deleted in . The Δ strain was also less virulent compared to wild type in an invasion model as reflected by a significant reduction in bacteria in the lungs and no dissemination to the blood and brain. Kinetic measurements with recombinant Har demonstrated that this enzyme reduced HOSCN with near diffusion-limited catalytic efficiency, using either NADH (k/K = 1.2 × 10 Ms) or NADPH (k/K = 2.5 × 10 Ms) as electron donors. We determined the X-ray crystal structure of Har in complex with the FAD cofactor to 1.50 Å resolution, highlighting the active site architecture characteristic for this class of enzymes. Collectively, our results demonstrate that pneumococcal Har is a highly efficient HOSCN reductase, enabling survival against oxidative host immune defenses. In addition, we provide structural insights that may aid the design of Har inhibitors.

中文翻译：

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次硫氰酸还原酶对于肺炎链球菌的宿主定植和感染至关重要

主要的人类病原体在定植和感染部位遇到免疫衍生的氧化剂次硫氰酸（HOSCN）。我们最近鉴定了肺炎球菌次硫氰酸还原酶 (Har)，它是黄素蛋白二硫键还原酶家族的成员，并表明它有助于 的 HOSCN 耐受。在这里，我们在肺炎球菌感染的小鼠模型中证明了 Har 对于定植和侵袭至关重要。在定植模型中，当 中删除时，鼻咽部的细菌负荷显着减弱。在入侵模型中，与野生型相比，Δ菌株的毒力也较低，这反映在肺部细菌显着减少并且没有传播到血液和大脑。重组 Har 的动力学测量表明，该酶使用 NADH (k/K = 1.2 × 10 Ms) 或 NADPH (k/K = 2.5 × 10 Ms) 作为电子供体，以接近扩散限制的催化效率还原 HOSCN。我们以 1.50 Å 的分辨率确定了 Har 与 FAD 辅因子复合物的 X 射线晶体结构，突出了此类酶的活性位点结构特征。总的来说，我们的结果表明肺炎球菌 Har 是一种高效的 HOSCN 还原酶，能够抵抗宿主氧化免疫防御。此外，我们提供的结构见解可能有助于 Har 抑制剂的设计。