Identification and Characterization of a Single High‐Affinity Fatty Acid Binding Site in Human Serum Albumin

Abstract

As ingle high-affinity fatty acid binding site in the important human transport protein serum albumin (HSA) is identified and characterized using an NBD (7-nitrobenz-2oxa-1,3-diazol-4-yl)-C 12 fatty acid. This ligand exhibits a1 :1 binding stoichiometry in its HSA complex with high sitespecificity.The complex dissociation constant is determined by titration experiments as well as radioactive equilibrium dialysis.C ompetition experiments with the knownH SA-binding drugs warfarin and ibuprofen confirm the new binding site to be different from Sudlow-sites Ia nd II. These binding studies are extended to other albumin binders and fatty acid derivatives.F urthermore an X-ray crystal structure allows locating the binding site in HSA subdomain IIA. The knowledge about this novel HSA site will be important for drug depot development and for understanding drug-protein interaction, which are important prerequisites for modulation of drug pharmacokinetics.

Cellular metabolic activity marker via selective turn-ON detection of serum albumin using an NBD-based fluoroprobe

“…This interesting result inspired us to investigate further the binding phenomena of NBD-Bu with BSA. As small molecules with similar van der Waals volume are known to bind with SA in 1:1 mode, 20 hence, in this case, the binding curve was fitted (Fig. 2b) using the with 1:1 binding equation.…”

“…19 Nitrobenzoxadiazole (NBD) dyes are well explored in the literature for their interesting fluorogenic intramolecular charge-transfer (ICT) properties and utilized for sensing, and protein binding studies. 20 NBD-labeled lipids are well-explored fluorescent probes for understanding membrane structure and dynamics, and have been widely used in both model systems and living cells. [21][22] Among the several hydrophobic binding cavities inside BSA, warfarin binds to site I while ibuprofen binds to site II.…”