Department of Biochemistry, Indian Institute of Science, Bangalore, India.
Molecular Biophysics Unit, Indian Institute of Science, Bangalore, India.
J Bacteriol. 2023 Jul 25;205(7):e0005923. doi: 10.1128/jb.00059-23. Epub 2023 Jun 27.
YciF (STM14_2092) is a member of the domain of unknown function (DUF892) family. It is an uncharacterized protein involved in stress responses in Salmonella Typhimurium. In this study, we investigated the significance of YciF and its DUF892 domain during bile and oxidative stress responses of Typhimurium. Purified wild-type YciF forms higher order oligomers, binds to iron, and displays ferroxidase activity. Studies on the site-specific mutants revealed that the ferroxidase activity of YciF is dependent on the two metal binding sites present within the DUF892 domain. Transcriptional analysis displayed that the ΔE strain, which has compromised expression of YciF, encounters iron toxicity due to dysregulation of iron homeostasis in the presence of bile. Utilizing this observation, we demonstrate that the bile mediated iron toxicity in ΔE causes lethality, primarily through the generation of reactive oxygen species (ROS). Expression of wild-type YciF, but not the three mutants of the DUF892 domain, in ΔE alleviate ROS in the presence of bile. Our results establish the role of YciF as a ferroxidase that can sequester excess iron in the cellular milieu to counter ROS-associated cell death. This is the first report of biochemical and functional characterization of a member of the DUF892 family. The DUF892 domain has a wide taxonomic distribution encompassing several bacterial pathogens. This domain belongs to the ferritin-like superfamily; however, it has not been biochemically and functionally characterized. This is the first report of characterization of a member of this family. In this study, we demonstrate that Typhimurium YciF is an iron binding protein with ferroxidase activity, which is dependent on the metal binding sites present within the DUF892 domain. YciF combats iron toxicity and oxidative damage caused due to exposure to bile. The functional characterization of YciF delineates the significance of the DUF892 domain in bacteria. In addition, our studies on Typhimurium bile stress response divulged the importance of comprehensive iron homeostasis and ROS in bacteria.
YciF（STM14_2092）是未知功能（DUF892）家族的一员。它是一种未被描述的蛋白质，参与沙门氏菌 Typhimurium 的应激反应。在这项研究中，我们研究了 YciF 及其 DUF892 结构域在 Typhimurium 胆汁和氧化应激反应中的重要性。纯化的野生型 YciF 形成更高阶的寡聚体，结合铁，并显示出亚铁氧化酶活性。对定点突变体的研究表明，YciF 的亚铁氧化酶活性依赖于 DUF892 结构域内存在的两个金属结合位点。转录分析显示，ΔE 菌株由于胆汁存在时铁稳态失调而导致铁毒性，该菌株的 YciF 表达受到损害。利用这一观察结果，我们证明了胆汁介导的 ΔE 中的铁毒性会导致细胞死亡，主要是通过产生活性氧（ROS）。在胆汁存在的情况下，野生型 YciF 的表达，但不是 DUF892 结构域的三个突变体的表达，可减轻 ΔE 中的 ROS。我们的结果确立了 YciF 作为一种亚铁氧化酶的作用，它可以在细胞环境中隔离多余的铁，以对抗与 ROS 相关的细胞死亡。这是首次对 DUF892 家族成员进行生化和功能表征的报道。DUF892 结构域具有广泛的分类学分布，包括几种细菌病原体。该结构域属于铁蛋白超家族；然而，它尚未在生化和功能上进行表征。这是首次对该家族成员进行特征描述的报道。在这项研究中，我们证明了沙门氏菌 Typhimurium 的 YciF 是一种具有亚铁氧化酶活性的铁结合蛋白，该活性依赖于 DUF892 结构域内存在的金属结合位点。YciF 可对抗由于暴露于胆汁而引起的铁毒性和氧化损伤。YciF 的功能表征描绘了 DUF892 结构域在细菌中的重要性。此外，我们对沙门氏菌胆汁应激反应的研究揭示了细菌中全面的铁稳态和 ROS 的重要性。
