Binding of Doxyl Stearic Spin Labels to Human Serum Albumin: An EPR Study

• Published in: The journal of physical chemistry. B 2014-09, Vol.118 (37), p.10898-10905
• Main Authors: Pavićević, Aleksandra A, Popović-Bijelić, Ana D, Mojović, Miloš D, Šušnjar, Snežana V, Bačić, Goran G
• Format: Article
• Language: English
• Subjects: Ascorbic Acid - chemistry; Cyclic N-Oxides - chemistry; Cyclic N-Oxides - metabolism; Electron Spin Resonance Spectroscopy; Ethanol - chemistry; Humans; Ibuprofen - chemistry; Protein Binding; Serum Albumin - chemistry; Serum Albumin - metabolism; Spin Labels; Temperature; Warfarin - chemistry
• ISSN: 1520-6106; 1520-5207
• DOI: 10.1021/jp5068928

The binding of spin-labeled fatty acids (SLFAs) to the human serum albumin (HSA) examined by electron paramagnetic resonance (EPR) spectroscopy was studied to evaluate the potential of the HSA/SLFA/EPR technique as a biomarking tool for cancer. A comparative study was performed on two spin labels with nitroxide groups attached at opposite ends of the fatty acid (FA) chain, 5-doxyl stearic (5-DS) and 16-doxyl stearic (16-DS) acid. The effects of incubation time, different [SLFA]/[HSA] molar ratios, ethanol, and temperature showed that the position of the nitroxide group produces certain differences in binding between the two SLFAs. Spectra for different [SLFA]/[HSA] molar ratios were decomposed into two spectral components, which correspond to the weakly and strongly bound SLFAs. The reduction of SLFA with ascorbate showed the existence of a two component process, fast and slow, confirming the decomposition results. Warfarin has no effect on the binding of the two SLFAs, whereas ibuprofen significantly decreases the binding of 5-DS and has no effect on 16-DS. Together, the results of this study indicate that both SLFAs, 5-DS and 16-DS, should be used for the study of HSA conformational changes in blood induced by various medical conditions.