Human serum albumin (HSA) is the most abundant protein in plasma and acts as one of the main carriers in the circulatory system. HSA participates in binding and transportation of a broad range of ligands such as fatty acids, drugs, hormones and other biologically important components that are essential for normal functioning of the body. HSA possesses multiple binding sites and is made up of three homologous domains (I, II and III) and each domain is made up of two subdomains (A and B) connected by random coil. There are a total of 9 binding pockets (FA1-FA9) with FA7, also named as Sudlow’s binding site, being the major drug binding site.
The majority of studies on HAS-ligand interactions are conducted using theoretical techniques such as classical molecular dynamics (MD) simulations and molecular docking. We have utilized the power of hydrogen deuterium exchange in combination with chemical cross-linking mass spectrometry approaches to perform epitope mapping at the molecular level. This study provides an important insight by mapping exact binding location of the drug ligand to HSA at near native conditions. We have demonstrated the capability of this approach which could help in new antibiotic designs with effective pharmacological properties.