Electrochemically nitrated equine myoglobin at tyrosine 103: The structural consequences of the role of NO2

Abstract

Myoglobin (Mb) is one of the most studied proteins and serves as model protein for evaluation and characterization the ligand–protein complexes such as the reversible binding of small molecules at the haem group. Nitration in proteins is a physiologically relevant process and the formation of 3-nitrotyrosine in vivo was first proposed as an in vivo marker of the production of reactive nitrogen species (RNS). We have performed the selectively nitration of Mb as model protein to carry out spectroscopic and electrochemical studies in order to add insights into the changes caused by nitration. The selection of the electrode, boron doped diamond (BDD), is critical as minimization of protein adsorption and retention of lytic activity is required to perform the electro-oxidative mononitration of Mb at Tyr103. Fig. 1 shows the voltammograms of the electrochemical reduction of native-Mb (solid line) at BDD electrode in sodium acetate buffer pH 4.0 in presence of saturated O2 conditions corresponds to the electrochemical reduction of MbFe (III) O2, which shows a slight decrease in the catalysis of O2 for NO2-Mb (dashed line). The study of the conformational and functional changes of the modified has been performed by the application of some spectroscopic techniques such as circular dichroism (CD), fluorescence, thermal stabilities and nuclear magnetic resonance. A relationship of thermal stability, structure and electrochemical behaviour of oxygen reduction has been observed for the electrosynthesis of the nitrated protein (NO2-Mb).