Department of Chemistry, The Pennsylvania State University, University Park, PA 16802.
Athinoula A. Martinos Center for Biomedical Imaging, Department of Radiology, Massachusetts General Hospital / Harvard Medical School, Charlestown, MA 02129.
Proc Natl Acad Sci U S A. 2022 Dec 20;119(51):e2212723119. doi: 10.1073/pnas.2212723119. Epub 2022 Dec 12.
The design of selective metal-binding sites is a challenge in both small-molecule and macromolecular chemistry. Selective recognition of manganese (II)-the first-row transition metal ion that tends to bind with the lowest affinity to ligands, as described by the Irving-Williams series-is particularly difficult. As a result, there is a dearth of chemical biology tools with which to study manganese physiology in live cells, which would advance understanding of photosynthesis, host-pathogen interactions, and neurobiology. Here we report the rational re-engineering of the lanthanide-binding protein, lanmodulin, into genetically encoded fluorescent sensors for Mn, MnLaMP1 and MnLaMP2. These sensors with effective (Mn) of 29 and 7 µM, respectively, defy the Irving-Williams series to selectively detect Mn in vitro and in vivo. We apply both sensors to visualize kinetics of bacterial labile manganese pools. Biophysical studies indicate the importance of coordinated solvent and hydrophobic interactions in the sensors' selectivity. Our results establish lanmodulin as a versatile scaffold for design of selective protein-based biosensors and chelators for metals beyond the f-block.
设计具有选择性的金属结合位点是小分子和生物大分子化学领域共同面临的挑战。如 Irving-Williams 序列所描述的，锰（II）——第一过渡金属离子，通常与配体的亲和力最低——的选择性识别尤其具有难度。因此，缺乏化学生物学工具来研究活细胞中的锰生理学，这将有助于推进对光合作用、宿主-病原体相互作用和神经生物学的理解。在这里，我们报告了对镧系元素结合蛋白 lanmodulin 的合理重新设计，将其构建成用于 Mn 的遗传编码荧光传感器 MnLaMP1 和 MnLaMP2。这两个传感器的有效浓度（Mn）分别为 29 和 7 µM，分别违背了 Irving-Williams 序列，能够在体外和体内选择性地检测 Mn。我们应用这两种传感器来可视化细菌不稳定锰库的动力学。生物物理研究表明，传感器选择性中的配位溶剂和疏水性相互作用非常重要。我们的研究结果确立了 lanmodulin 作为一种多功能支架，可用于设计针对除 f 区以外的金属的具有选择性的基于蛋白质的生物传感器和螯合剂。
