Mapping the Conformational Dynamics of E-selectin upon Interaction with its Ligands

• Main Author: Aleisa, Fajr A
• Other Authors: Merzaban, Jasmeen
• Language: en
• Published: 2013
• Subjects: E-Selectin; cloning; Sf9 cells; CHO-Kl cells; Hermatopoietic stem cells (HSC); Forster resonance energy transfer (FRET)
• Online Access: Aleisa, F. A. (2013). Mapping the Conformational Dynamics of E-selectin upon Interaction with its Ligands. KAUST Research Repository. https://doi.org/10.25781/KAUST-YY239; http://hdl.handle.net/10754/292320

Selectins are key adhesion molecules responsible for initiating a multistep process that leads a cell out of the blood circulation and into a tissue or organ. The adhesion of cells (expressing ligands) to the endothelium (expressing the selectin i.e.,E-selectin) occurs through spatio-temporally regulated interactions that are mediated by multiple intra- and inter-cellular components. The mechanism of cell adhesion is investigated primarily using ensemble-based experiments, which provides indirect information about how individual molecules work in such a complex system. Recent developments in single-molecule (SM) fluorescence detection allow for the visualization of individual molecules with a good spatio-temporal resolution nanometer spatial resolution and millisecond time resolution). Furthermore, advanced SM fluorescence techniques such as Förster Resonance Energy Transfer (FRET) and super-resolution microscopy provide unique opportunities to obtain information about nanometer-scale conformational dynamics of proteins as well as nano-scale architectures of biological samples. Therefore, the state-of-the-art SM techniques are powerful tools for investigating complex biological system such as the mechanism of cell adhesion. In this project, several constructs of fluorescently labeled E-selectin will be used to study the conformational dynamics of E-selectin binding to its ligand(s) using SM-FRET and combination of SM-FRET and force microscopy. These studies will be beneficial to fully understand the mechanistic details of cell adhesion and migration of cells using the established model system of hematopoietic stem cells (HSCs) adhesion to the selectin expressing endothelial cells (such as the E-selectin expressing endothelial cells in the bone marrow).