title: Reevaluation of ANS binding to human and bovine serum albumins: key role of equilibrium microdialysis in ligand - receptor binding characterization.
abstract: In this work we return to the problem of the determination of ligand-receptor binding stoichiometry and binding constants. In many cases the ligand is a fluorescent dye which has low fluorescence quantum yield in free state but forms highly fluorescent complex with target receptor. That is why many researchers use dye fluorescence for determination of its binding parameters with receptor, but they leave out of account that fluorescence intensity is proportional to the part of the light absorbed by the solution rather than to the concentration of bound dye. We showed how ligand-receptor binding parameters can be determined by spectrophotometry of the solutions prepared by equilibrium microdialysis. We determined the binding parameters of ANS - human serum albumin (HSA) and ANS - bovine serum albumin (BSA) interaction, absorption spectra, concentration and molar extinction coefficient, as well as fluorescence quantum yield of the bound dye. It was found that HSA and BSA have two binding modes with significantly different affinity to ANS. Correct determination of the binding parameters of ligand-receptor interaction is important for fundamental investigations and practical aspects of molecule medicine and pharmaceutics. The data obtained for albumins are important in connection with their role as drugs transporters.
authors: Laboratory of Structural Dynamics, Stability and Folding of Proteins of the Institute of Cytology, Russian Academy of Sciences, St. Petersburg, Russian Federation.
authors: Irina M
authors: IM
authors: Kuznetsova
authors: Anna I
authors: AI
authors: Sulatskaya
authors: Olga I
authors: OI
authors: Povarova
authors: Konstantin K
authors: KK
authors: Turoverov
mesh_terms: D000817:Anilino Naphthalenesulfonates / Q000378:metabolism*; D000818:Animals; D002417:Cattle; D006801:Humans; D017551:Microdialysis; D011485:Protein Binding; D012710:Serum Albumin, Bovine / Q000378:metabolism*
chemical_list: D000817:Anilino Naphthalenesulfonates; D012710:Serum Albumin, Bovine; C027132:1-anilino-8-naphthalenesulfonate