IRON TRANSPORT, SYNERGISTIC ANIONS

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。我们正在研究铁摄取系统的机制，该系统对于致病性金黄色葡萄球菌在铁限制的介质中的生存至关重要。异羟甲酸铁摄取系统能够识别和同化异羟甲酸铁的铁载体；它由一个完整的膜ABC转运蛋白复合体和外周结合蛋白FhuD1和FhuD2组成。我们对铁铁载体结合蛋白特别感兴趣，因为它们对结构多样化的铁载体具有很高的亲和力，可以用来以“特洛伊木马”的方式向细菌输送抗生素。在Beamiline ID18上使用SAXS，我们证明了FhuD1和FhuD2在与配体结合时只发生了很小的构象变化。事实上，最近的测量表明，构象变化的程度取决于铁载体的确切性质。在我们的出版物中讨论了铁载体和结合蛋白在运输机制中的作用。重要的是，SAXS数据提供的结构信息使我们得出结论，ABC运输复合体对配体的识别是Fhu系统运输机制的关键部分。我们还在E.Coli中使用麦芽糖结合蛋白(MBP)的构象工程来评估结合蛋白和配体在通过完整的膜复合体触发ATP水解中的相对作用。在光束线ID18获得的SAXS数据对于快速准确地评估我们的突变对MBP构象的影响是绝对关键的。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. We are investigating the mechanism of an iron uptake system that is important for survival of pathogenic Staphylococcus aureus in iron-limited media. The Ferric Hydroxamate Uptake ("Fhu") system is able to recognize and assimilate ferric hydroxamate siderophores; it is comprised of an integral membrane ABC transporter complex and peripheral binding proteins, FhuD1 and FhuD2. We are particularly interested in the ferric siderophore binding proteins because they have a high affinity for a structurally diverse collection of siderophores and could be used to deliver antibiotics to the bacteria in a "Trojan Horse" manner. Using SAXS at beamiline ID18, we demonstrated that FhuD1 and FhuD2 undergo only a very small conformational change when they bind ligand. In fact, recent measurements indicate that the extent of the conformational change depends on the exact nature of the siderophore. The role of the siderophore and the binding protein in the transport mechanism are discussed in our publications. Importantly, the structural information provided by SAXS data has led us to conclude that recognition of the ligand by the ABC transport complex is a critical part of the transport mechanism for the Fhu system. We are also using conformational engineering of maltose binding protein (MBP) in E. Coli to assess the relative roles of the binding protein and ligand in triggering ATP hydrolysis by the integral membrane complex. SAXS data obtained at beamline ID18 have been absolutely critical for rapid and accurate evaluation of the effect of our mutations on the conformation of MBP.