Scavenging of hydrated electron by HSA or Ligand/HSA adduct: Pulse radiolysis study

Abstract

The process of electron scavenging by molecule of human serum albumin in solution under neutral pH was studied. The rate constant of reaction eaq- with HSA is diffusion controlled (1.1 × 1010 dm3 mol−1). The transient absorption spectrum recorded during pulse radiolysis of HSA solution under reductive conditions shows maximum at 420 nm. The hydrated electron may react with many targets within albumin. We conducted a series of pulse radiolysis experiments with aqueous solutions containing amino acids, including: cysteine, cystine, L-tyrosine, L-tryptophan, methionine, dimer tyrosine- alanine and dimer alanine-tryptophan. The analysis of the shape of transient spectra and the reactivity of e-aq with amino acids and HSA suggest that electron attachment to disulfide bond (CyS-SCy•−) is responsible for transient absorption spectrum recorded in the case of protein solution. The molecule of HSA did not undergo significant size changes after irradiation but molecular damage at Sudlow's site I (subdomain IIA) is detected and proved by changes in fluorescence properties of Trp214. The breaking of S–S bonds, and the desulfurization some of them, may be connected with the structural modification of HSA.The decay process of CyS-SCy•− is clearly detected during pulse radiolysis of HSA solution above 329 K. The influence of HSA on electron scavenging by selected ligands (9,10-anthraquinone-1,5-disulfonate (AQDS2-), 1- pyrene sulfonate (PSA-), rose bengal (RB2-), methylene blue (MB+), indocyanine green (ICG-)) was studied. The solute molecules (ligands) embedded in a hydrophobic domain at Sudlow's site 1 (AQDS2-, PSA-, RB2-) are very well protected against eaq- attack. The methylene blue localizes predominantly in the protein binding site 2 (subdomain 3A). In contrast to site 1, the entrance to site 2 is exposed to the solvent and MB+ molecule is located within HSA structure in space accessible for e-aq. As a consequence, MB+ is relatively easy reduced by hydrated electron.