A distal ligand mutes the interaction of hydrogen sulfide with human neuroglobin

Abstract

Edited by Norma AllewellHydrogen sulfide is a critical signaling molecule, but high concentrations cause cellular toxicity. A four-enzyme pathway in the mitochondrion detoxifies H 2 S by converting it to thiosulfate and sulfate. Recent studies have shown that globins like hemoglobin and myoglobin can also oxidize H 2 S to thiosulfate and hydropolysulfides. Neuroglobin, a globin enriched in the brain, was reported to bind H 2 S tightly and was postulated to play a role in modulating neuronal sensitivity to H 2 S in conditions such as stroke. However, the H 2 S reactivity of the coordinately saturated heme in neuroglobin is expected a priori to be substantially lower than that of the 5-coordinate hemes present in myoglobin and hemoglobin. To resolve this discrepancy, we explored the role of the distal histidine residue in muting the reactivity of human neuroglobin toward H 2 S. Ferric neuroglobin is slowly reduced by H 2 S and catalyzes its inefficient oxidative conversion to thiosulfate. Mutation of the distal His 64 residue to alanine promotes rapid binding of H 2 S and its efficient conversion to oxidized products. X-ray absorption, EPR, and resonance Raman spectroscopy highlight the chemically different reaction options influenced by the distal histidine ligand. This study provides mechanistic insights into how the distal heme ligand in neuroglobin caps its reactivity toward H 2 S and identifies by cryo-mass spectrometry a range of sulfide oxidation products with 2-6 catenated sulfur atoms with or without oxygen insertion, which accumulate in the absence of the His 64 ligand.Neuroglobin is a member of the globin family (1) that is expressed primarily in neuronal tissues (2) but also in other metabolically highly active endocrine tissues, such as the adrenal gland and the pancreatic islets of Langerhans (3, 4). Although the physiological role of neuroglobin is still elusive, early studies suggested that it might be involved in oxygen supply (1, 5). However, neuroglobin is generally expressed at low levels and exhibits a high autoxidation rate, producing reactive oxygen species, which makes its role as an O 2 carrier unlikely (6). Neuroglobin can also scavenge reactive oxygen species and reduce nitrite to NO under hypoxic conditions (7-9).In contrast to hemoglobin and myoglobin, two histidine residues coordinate the heme cofactor in neuroglobin in the ferrous and ferric states (Fig. 1A). The crystal structure of neuroglobin reveals a high structural similarity to myoglobin, despite Ͻ25% amino acid sequence similarity between the proteins. The structure reveals a large hydrophobic cavity, which connects the proximal and distal sides of the heme (7). His 64 and His 96 coordinate the heme on the distal and proximal sides, respectively (10). Exogenous ligands like O 2 , CO, and NO bind to ferrous neuroglobin (Fe II -Ngb) 2 on the distal side, and binding is limited by dissociation of His 64 , which is slow (6,(11)(12)(13)(14). Limited characterization of the binding of sulfide to ferric neuroglobin (Fe III -Ngb) has...

International Union of Basic and Clinical Pharmacology. CII: Pharmacological Modulation of H2S Levels: H2S Donors and H2S Biosynthesis Inhibitors

“…Although scavengers have many inherent problems (specificity, selectivity, delivery issues, and, unless they are catalytic, the fact they are consumed in the reactions and typically require large concentrations/ doses), in theory, this approach may also be of some merit for further exploration. However, the current state-of-the-art of H 2 S scavengers is in an embryonic stage; although heme-containing proteins (e.g., hemoglobin, myoglobin, neuroglobin) are known to scavenge H 2 S (Brunyanszki et al, 2015;Bostelaar et al, 2016;Ruetz et al, 2017;Vitvitsky et al, 2017), they also scavenge many other reactive species including nitric oxide.…”

Crystal structure of Kumaglobin: a hexacoordinated heme protein from an anhydrobiotic tardigrade, Ramazzottius varieornatus

“…A). Previously reported EPR spectroscopic data demonstrates that human Ngb shows high‐spin ferric signals as well as low‐spin ones because of dissociation of distal histidine from the iron atom in the presence of disulfide bond between Cys46 and Cys55 . Unlike human Ngb, the EPR spectrum of WT Kgb did not show any sign of high‐spin signals, implying that the two axial ligands in Kgb are tightly bound to the iron atom.…”

“…However, while gaseous ligands are bound at the distal position through a heme sliding mechanism in Ngb and a distal histidine flipping mechanism in Cgb , it is unlikely that Kgb is responsible for binding gaseous ligands. Although dissociation of distal histidine from iron in Ngb is supported by the formation of a disulfide bond between Cys46 and Cys55 , such pair forming disulfide bond is absent in Kgb. Therefore, if Kgb reacts with oxygen species, it will likely undergo an outer‐sphere mechanism in which a heme–O 2 complex is not formed , while human Ngb reacts by forming a heme–O 2 complex to scavenge superoxide .…”

“…The mutant showed peaks of the Soret band and the b band at 405 and around 500 nm, respectively. A slight blue shift of the Soret band and weak absorption between 600 and 650 nm indicated that H107A Kgb had pentacoordinated heme [38,46,48]. Upon adding H 2 S, a peak shift from 405 to 424 nm was observed in the spectrum of the mutant.…”

Mechanism-based inhibition of human persulfide dioxygenase by γ-glutamyl-homocysteinyl-glycine

“…Alternatively or additionally, increased H 2 S could stimulate its oxidation by heme proteins. We have shown that globins like hemoglobin, myoglobin and neuroglobin catalyze oxidation of H 2 S to thiosulfate (26)(27)(28)(29).…”