The use of trinitrobenzenesulfonic acid in studies on the binding of fatty acid anions to bovine serum albumin

The kinetics of the reaction between trinitrobenzenesulfonic acid (TNBS) and bovine serum albumin (BSA) have been studied. The results obtained indicate that TNBS is bound to certain binding sites on BSA and reacts with amino groups present there. The presence of palmitate strongly affects the rate of the reaction between TNBS and BSA probably by competition for the same binding sites. Tryptic digestion of BSA labeled with TNBS in the presence of various amounts of palmitate followed by fractionation of the peptides formed showed that labeling of one of the peptides was strongly inhibited by the presence of palmitate. This peptide was isolated and further digested with chymotrypsin yielding a labeled pentapeptide. The sequence was determined and showed a tyrosine residue next to the labeled lysine.The binding affinity of caprylate and palmitate to native and TNBS-treated BSA was studied by equilibrium dialysis. A model for the binding of fatty acid anions to BSA is suggested.