The effect of aspirin-HSA complexation on the protein secondary structure

This study was designed to determine the secondary structure of human serum albumin (HSA) in the presence of aspirin in H2O and D2O solutions at physi ological pH, using aspirin concentrations of 0.0001-5 mM with final protein concentration of 2% w/v. UV-vis spectra and Fourier transform infrared (FT IR) difference spectroscopy with its self-deconvolution, second derivative resolution enhancement, and curve-fitting procedures were applied to charac terize the drug binding mode, the binding constant, and the protein seconda ry structure in the aspirin-HSA complexes. Spectroscopic evidence showed th at no aspirin-protein interaction occurs at very low drug concentration (0. 0001 mM), whereas at higher drug contents (0.001-0.1 mM) the aspirin anion binding (H-bonding) is mainly through the epsilon-amino NH3+ group with ove rall binding constant of K = 1.4 x 10(4) M-1. At high drug concentrations ( 1-5 mM), acetylation of Lys-199 was observed. Aspirin binding results in pr otein secondary structural changes from that of the alpha-helix 55% (free H SA) to 49%, beta-sheet 22% (free HSA) to 31%, beta-anti 12% (free HSA) to 4 % and turn 11% (free HSA) to 16% in the aspirin-HSA complexes.