Thiol-disulfide exchange reactions occurring at modified bovine serum albumin detected using ellman's reagent (5, 5'-dithiobis (2-itrobenzoic acid)

• Published in: Pakistan journal of pharmaceutical sciences Vol. 33; no. 6(Supplementary); pp. 2767 - 2772
• Main Authors: Khan, Haroon, Khan, Kamran Ahmad, Khan, Muhammad Khalid, Ahmad, Ashfaq, Niazi, Zahid Rasul, Mangi, Altaf, -Rehman, Fazal-Ur, Shah, Kifayat Ullah, Gul, Rahman, Jan, Syed Umer
• Format: Journal Article
• Language: English
• Published: Pakistan Pakistan Journal of Pharmaceutical Sciences 01.11.2020
• Subjects: Acetylcysteine; Albumin; Analysis; Chemical tests and reagents; Cystine; Mediation; Spectrum analysis; Thiols; Pakistan
• ISSN: 1011-601X
• DOI: 10.36721/PJPS.2020.33.6.SUP.2767-2772.1

Bovine serum albumin (BSA) is usually employed as a model protein because of being homologous with human serum albumin. Cysteine-34 of BSA has been oxidised with Ellman's reagent to produce BSA labelled with an Ellman's moiety (BSA-SE). The BSA-SE was then reacted with glutathione, N-acetylcysteine and D-penicillamine (D-pen). The two were able to release the Ellman's moiety bound at cysteine-34 while D-pen did not. Albumin labeled using Ellman's reagent was used to demonstrate the cleavage of a protein mixed disulphide. The kinetics of thiol disulfide interchange reactions involving formation of a chromophoric thiolate were determined by UV-visible spectroscopy. The reaction of thiolates with excess Ellman's reagent is used for quantitative estimation of thiol by measuring the absorption at λ, 412 nm. The disulfide exchange reactions occurring at Cys-34 of BSA was determined and the reduction of oxidized Cys-34 was studied in order to understand the reverse reaction. Spectroscopic evidence suggested that glutathione and N-acetylcysteine remove the label and produce BSA in a disulfide form. In contrast, D-pen reaction returned BSA to its thiolate form via mediation. It was observed that thio-disulfide exchange occurred at cysteine-34 labelled with Ellman's moiety. The implications to the redox status of plasma are discussed.