A spectroscopic and kinetic study of the singlet oxygen mediated oxidation of human and bovine serum albumin

DORADO RD; ESPECHE TURBAY, MB.; BORSARELLI, CD.

La Plata

Congreso; XLVII Reunión Anual SAB; 2018

Singlet oxygen (1O2) is a very harmful non-radical species that reacts withelectron-rich biological molecules. Due to their intrinsic abundance in thebiological milieu, proteins are the main targets of action of 1O2, causing structurechanges, covalent and non-covalent aggregation, and enzymatic activity losses.Serum albumins (SAs), are fundamental for the maintenance of the oncoticpressure of the plasma and the transporting of small vital molecules and drugs.As well as the functionality of both SA are depending on the protein structure, inthis report we evaluated the impact of the 1O2 oxidation on the human (HSA) andbovine (BSA) SAs, which show an amino acid similarity of about 77%, but differingmainly in the number of Trp and Tyr residues, two of the oxidizable residues by 1O2besides Cys, Met and His. The ruthenium (II) tris(2,2'-bipyridine) cation (Rubpy, 20μM) was used as photosensitizer for generation of 1O2, since Rubpy does not bindto the SAs (10-100 μM). Photolysis was performed using a 1 W blue LED (443±27nm) and the oxidation kinetics was monitored by both absorption and fluorescencespectroscopy. Post-irradiation the sensitizer was separated by ion exchangechromatography, and the modified proteins were analyzed by severalspectroscopies, laser particle electrophoresis (Z potential), protein electrophoresis(PAGE); carbonyl content and pseudo-esterase activity determination throughbiochemical techniques. Results indicated the oxidative degradation of Trpresidues with the production of carbonylic fluorescent products. The primarystructure of the SA was retained, despite changes of the superficial electricalpotential and hydrophobicity, and the reduction of the pseudo-esterase activity.Albeit both SAs share similar globular structure and amino acid sequence, thepresent results indicates subtle differences in the impact of the 1O2-mediatedprotein oxidation.