Department of Microbiology and Immunology, Life Sciences Institute, The University of British Columbia , Vancouver, BC, Canada V6T 1Z3.
Biochemistry. 2014 Apr 15;53(14):2286-94. doi: 10.1021/bi500230f. Epub 2014 Apr 1.
The Isd (iron-regulated surface determinant) system is a multiprotein transporter that allows bacterium Staphylococcus aureus to take up iron from hemoglobin (Hb) during human infection. In this system, IsdB is a cell wall-anchored surface protein that contains two near iron transporter (NEAT) domains, one of which binds heme. IsdB rapidly extracts heme from Hb and transfers it to IsdA for relay into the bacterial cell. Using a series of recombinant IsdB constructs that included at least one NEAT domain, we demonstrated that both domains are required to bind Hb with high affinity (KD = 0.42 ± 0.05 μM) and to extract heme from Hb. Moreover, IsdB extracted heme only from oxidized metHb, although it also bound oxyHb and the Hb-CO complex. In a reconstituted model of the biological heme relay pathway, IsdB catalyzed the transfer of heme from metHb to IsdA with a Km for metHb of 0.75 ± 0.07 μN and a kcat of 0.22 ± 0.01 s(-1). The latter is consistent with the transfer of heme from metHb to IsdB being the rate-limiting step. With both NEAT domains and the linker region present in a single contiguous polypeptide, high-affinity Hb binding was achieved, rapid heme uptake was observed, and multiple turnovers of heme extraction from metHb and transfer to IsdA were conducted, representing all known Hb-heme uptake functions of the full-length IsdB protein.
Isd（铁调节表面决定簇）系统是一种多蛋白转运体，使金黄色葡萄球菌能够在人体感染期间从血红蛋白（Hb）中摄取铁。在该系统中，IsdB 是一种细胞壁锚定的表面蛋白，包含两个近铁转运体（NEAT）结构域，其中一个结构域结合血红素。IsdB 从 Hb 中快速提取血红素并将其转移到 IsdA 中，以便进入细菌细胞。使用一系列包含至少一个 NEAT 结构域的重组 IsdB 构建体，我们证明了这两个结构域都需要以高亲和力（KD=0.42±0.05μM）结合 Hb，并从 Hb 中提取血红素。此外，IsdB 仅从氧化的 metHb 中提取血红素，尽管它也结合 oxyHb 和 Hb-CO 复合物。在生物血红素传递途径的重建模型中，IsdB 以 Km 为 0.75±0.07μN 的 metHb 催化血红素向 IsdA 的转移，kcat 为 0.22±0.01s(-1)。后者与 metHb 向 IsdB 的血红素转移是限速步骤一致。在单个连续多肽中同时存在两个 NEAT 结构域和连接区，实现了高亲和力的 Hb 结合，观察到快速的血红素摄取，并且从 metHb 中提取血红素并向 IsdA 转移进行了多次周转，代表全长 IsdB 蛋白的所有已知 Hb-血红素摄取功能。
