Center for Biomolecular Structure Lecture Series

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X-ray Fluorescence Microscopy (XFM) is a powerful method for imaging the trace-element concentration, distribution, and speciation in cells and tissues. Even though the technique has been around for more than 30 years, only recently have advances in X-ray sources, optics, and detectors enabled two- and three-dimensional X-ray imaging at the nanoscale with attogram detection sensitivity. However, one limitation of XFM for imaging biological systems is detecting the trace-element distribution in the context of subcellular organelles and individual proteins. For visible light microscopy, the most ubiquitous method for imaging individual proteins within the context of a living cell is the use of intrinsically fluorescent proteins, such as the green fluorescent protein (GFP) that is co-expressed as a fusion tag along with the protein of interest. However, visualization of these tags is limited by the wavelengths of visible light except by applying specialized super resolution approaches. Here, we are developing applications enabled by encoded lanthanide-binding tags (LBTs), which are GFP-like analogs of minimal size (ca. 15-20 amino acids) for XFM. In this talk, applications of LBTs to both membrane-bound and cytosolic proteins will be demonstrated in 2D and 3D using a 15 nm X-ray beam. This approach enables visualization of LBT-tagged proteins while simultaneously measuring the elemental distribution in cells at a spatial resolution necessary for visualizing cell membranes and eukaryotic subcellular organelles.