Reactivity and endogenous modification by nitrite and hydrogen peroxide: does human neuroglobin act only as a scavenger? Available to Purchase

NGB (human neuroglobin), a recently discovered haem protein of the globin family containing a six-co-ordinated haem, is expressed in nervous tissue, but the physiological function of NGB is currently unknown. As well as playing a role in neuronal O₂ homoeostasis, NGB is thought to act as a scavenger of reactive species. In the present study, we report on the reactivity of metNGB (ferric-NGB), which accumulates in vivo as a result of the reaction of oxyNGB (oxygenated NGB) with NO, towards NO₂⁻ and H₂O₂. NO₂⁻ co-ordination of the haem group accounts for the activity of metNGB in the nitration of phenolic substrates. The two different metNGB forms, with and without the internal disulfide bond between Cys⁴⁶ (seventh residue on the inter-helix region between helices C and D) and Cys⁵⁵ (fifth residue on helix D), exhibit different reactivity, the former being more efficient in activating NO₂⁻. The kinetics of the reactions, the NO₂⁻-binding studies and the analysis of the nitrated products from different substrates all support the hypothesis that metNGB is able to generate an active species with the chemical properties of peroxynitrite, at pathophysiological concentrations of NO₂⁻ and H₂O₂. Without external substrates, the targets of the reactive species generated by the metNGB/NO₂⁻/H₂O₂ system are endogenous tyrosine (resulting in the production of 3-nitrotyrosine) and cysteine (oxidized to sulfinic acid and sulfonic acid) residues. These endogenous modifications were characterized by HPLC-MS/MS (tandem MS) analysis of metNGB after reaction with NO₂⁻ and H₂O₂ under various conditions. The internal S–S bond affects the functional properties of the protein. Therefore metNGB acts not only as scavenger of toxic species, but also as a target of the self-generated reactive species. Self-modification of the protein may be related to or inhibit its postulated neuroprotective activity.