Electrochemical nitration of myoglobin at tyrosine 103: Structure and stability

Evaluation of structural and stability changes of nitrated myoglobin at tyrosine 103 residues induced by electrochemical nitration. [Display omitted] ► Structural aspects of electrochemically-nitrated myoglobin. ► NMR data indicate that nitrated Mb has a slightly open conformation. ► Electrochemical nitration of equine skeletal muscle at amino acid tyrosine 103. ► Evaluation and characterization of structural and thermal stability changes. ► Subtle changes were observed involving tertiary structure slightly lost at pH 7. Nitration in proteins is a physiologically relevant process and the formation of 3-nitrotyrosine was first proposed as an in vivo marker of the production of reactive nitrogen species in oxidative stress. No studies have been published on structural changes associated with nitration of myoglobin. To address this deficiency the electrochemical nitration of equine skeletal muscle (Mb) at amino acid tyrosine 103 has been investigated for the evaluation and characterization of structural and thermal stability changes. Y103 in Mb is one of the most exposed tyrosine residues and it is also close to the heme group. Effects of Y103 nitration on the secondary and tertiary structure of Y103 have been studied by UV–Vis, circular dichroism, fluorescence and NMR spectroscopy and by electrochemical studies. At physiological pH, subtle changes were observed involving slight loosening of the tertiary structure and conformational exchange processes. Thermal stability of the nitrated protein was found to be reduced by 5°C for the nitrated Mb compared with the native Mb at physiological pH. Altogether, NMR data indicates that nitrated Mb has a very similar tertiary structure to that of native Mb, although with a slightly open conformation.