X‐band ESEEM spectroscopy of <sup>15</sup>N substituted native and inhibitor‐bound superoxide dismutase

Dikanov, Sergei A.; Felli, Isabella C.; Viezzoli, Maria Silvia; Spoyalov, Andrei P.; Hüttermann, Jürgen

doi:10.1016/0014-5793(94)00406-4

Cited by 10 publications

(7 citation statements)

References 20 publications

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“…The HYSCORE spectrum of SOD at g = 2.06 (Fig. 4B) shows two of these double quantum, dq-dq, cross peaks of nearly equal intensity that we can assign to a stronger set of hyperfine couplings due to H44 and H46, and a weaker set of couplings arising from H61 and H118 based on previous ESEEM studies (27). Treatment of the enzyme with H 2 O 2 causes the ESEEM depth or amplitude to decrease by about 40% and the dq-dq correlation in the HYSCORE spectrum (Fig.…”

Section: Discussionsupporting

confidence: 64%

Oxidation of Histidine Residues in Copper−Zinc Superoxide Dismutase by Bicarbonate-Stimulated Peroxidase and Thiol Oxidase Activities: Pulse EPR and NMR Studies

et al. 2010

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In this work, we investigated the oxidative modification of histidine residues induced by peroxidase and thiol oxidase activities of bovine copper-zinc superoxide dismutase (Cu-ZnSOD) using NMR and pulse EPR spectroscopy. 1D NMR and 2D-NOESY were used to determine the oxidative damage at the Zn(II) and Cu(II) active sites as well as at distant histidines. Results indicate that during treatment of SOD with hydrogen peroxide (H2O2) or cysteine in the absence of bicarbonate anion (HCO3−), both exchangeable and non-exchangeable protons were affected. Both His-44 and His-46 in the Cu(II) active site were oxidized based on the disappearance of NOESY cross peaks between CH and NH resonances of the imidazole rings. In the Zn(II) site, only His-69, which is closer to His-44, was oxidatively modified. However, addition of HCO3− protected the active site His residues. Instead, resonances assigned to His-41 residue, 11 Å away from the Cu(II) site, were completely abolished during both HCO3− stimulated peroxidase activity and thiol oxidase activity in the presence of HCO3−. Additionally, ESEEM/HYSCORE and ENDOR studies of SOD treated with peroxide/Cys in the absence of HCO3− revealed that hyperfine couplings to the distal and directly coordinated nitrogens of the His-44 and His-46 ligands at the Cu(II) active site were modified. In the presence of HCO3−, these modifications were absent. HCO3− mediated, selective oxidative modification of histidines in SOD may be relevant to understanding the molecular mechanism of SOD peroxidase and thiol oxidase activities.

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Section: Discussionsupporting

confidence: 64%

Oxidation of Histidine Residues in Copper−Zinc Superoxide Dismutase by Bicarbonate-Stimulated Peroxidase and Thiol Oxidase Activities: Pulse EPR and NMR Studies

et al. 2010

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“…It may be that the weak coupling of 0.42 MHz is mostly isotropic, and that the larger 0.8 MHz coupling is due to a second nitrogen. The 0.42 MHz splitting may be assigned to the distal nitrogen of the imidazole ring, as the hyperfine interaction is approximately 1/20th of that of the bound nitrogen, a common condition seen in imidizole-bound copper and vanadium systems and in Rieske clusters 51,56,62. More than one nitrogen are also visible in the X-band ESEEM data taken on 14 N-mitoNEET resulting in a very congested low frequency region where transitions would be expected to overlap (Fig.…”

Section: Resultsmentioning

confidence: 97%

Binding of Histidine in the (Cys)₃(His)₁-Coordinated [2Fe−2S] Cluster of Human mitoNEET

et al. 2010

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Human mitoNEET is a homodimeric iron-sulfur protein located in the outer mitochondrial membrane with unknown function, but which is known to interact with thiazolidinedione diabetes drugs. Each monomer houses a [2Fe-2S] cluster with an unusual (Cys) 3 (His) 1 ligation. The His ligand is important for enabling cluster release and for tuning the redox potential. We use multi-frequency (X-, Ka-, Qband) and multi-technique (CW, ESEEM, ENDOR, HYSCORE) EPR spectroscopy to investigate the cluster in its paramagnetic reduced [Fe 2+ Fe 3+ ] (S = 1/2) state. It has a rhombic g-tensor (2.007, 1.937, 1.897) with an average g-value of 1.947 that falls between those of Rieske-type and ferredoxintype [2Fe-2S] clusters. Simulation and least-squares fitting of orientation-selective Ka-and Q-band ENDOR, 1D ESEEM and HYSCORE spectra of 14 N and 15 N-labelled mitoNEET yield the principal values and orientations of both the hyperfine tensor ( 14 N: A iso = −6.25 MHz, T = −0.94 MHz) and the quadrupolar tensor (e 2 Qq/h = −2.47 MHz, η = +0.38) of the ligating histidine nitrogen N δ . From these, we can infer the absolute g tensor orientation with respect to the cluster: The g 2 axis is close to perpendicular to the [2Fe-2S] plane, and g 1 and g 2 are in-plane, but skewed from the Fe-Fe and S-S axes. In X-band ENDOR and ESEEM spectra, a weakly coupled nitrogen is visible, most likely the N ε of the histidine in the protonated state. We find that the cluster is in a valence-localized state, where Fe 2+ is His-bound. The field-sweep spectra show evidence of inter-cluster dipolar coupling that can be simulated using an uncoupled spin model for each cluster (S Fe2+ = 2, S Fe3+ = 5/2). The parameters determined in this work can function as reporters on how the cluster structure is altered upon pH changes and drug binding.

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“…How such a subtle change would account for different EPR signals is not clear, so the two copies are assumed not to account for the two very similar, but distinguishable, type 2 Cu 2ϩ signals. If the Cu 2ϩ binding site is formed from three imidazoles or if the imidazoles are inequivalent (for example, ␦ and nitrogen donor atoms), cis and trans configurations or different orientations of the imidazole rings from histidine (for example, rings perpendicular or parallel to the square plane of the complex; Dikanov et al, 1994a) might account for two very similar type 2 Cu 2ϩ sites.…”

Section: Possible Binding Sites For Type 2 Cu 2؉ In Pmmomentioning

confidence: 99%

Comparison of EPR-Visible Cu2+ Sites in pMMO from Methylococcus capsulatus (Bath) and Methylomicrobium album BG8

Lemos

Collins

Eaton

et al. 2000

Biophysical Journal

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X-band (9.1 GHz) and S-band (3.4 GHz) electron paramagnetic resonance (EPR) spectra for particulate methane monooxygenase (pMMO) in whole cells from Methylococcus capsulatus (Bath) grown on (63)Cu and (15)N were obtained and compared with previously reported spectra for pMMO from Methylomicrobium album BG8. For both M. capsulatus (Bath) and M. album BG8, two nearly identical Cu(2+) EPR signals with resolved hyperfine coupling to four nitrogens are observed. The EPR parameters for pMMO from M. capsulatus (Bath) (g( parallel) = 2.244, A( parallel) = 185 G, and A(N) = 19 G for signal one; g( parallel) = 2.246, A( parallel) = 180 G, and A(N) = 19 G for signal two) and for pMMO from M. album BG8 (g( parallel) = 2.243, A( parallel) = 180 G, and A(N) = 18 G for signal one; g( parallel) = 2. 251, A( parallel) = 180 G, and A(N) = 18 G for signal two) are very similar and are characteristic of type 2 Cu(2+) in a square planar or square pyramidal geometry. In three-pulse electron spin echo envelope modulation (ESEEM) data for natural-abundance samples, nitrogen quadrupolar frequencies due to the distant nitrogens of coordinated histidine imidazoles were observed. The intensities of the quadrupolar combination bands indicate that there are three or four coordinated imidazoles, which implies that most, if not all, of the coordinated nitrogens detected in the continuous wave spectra are from histidine imidazoles.

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X‐band ESEEM spectroscopy of ¹⁵N substituted native and inhibitor‐bound superoxide dismutase

Cited by 10 publications

References 20 publications

Oxidation of Histidine Residues in Copper−Zinc Superoxide Dismutase by Bicarbonate-Stimulated Peroxidase and Thiol Oxidase Activities: Pulse EPR and NMR Studies

Oxidation of Histidine Residues in Copper−Zinc Superoxide Dismutase by Bicarbonate-Stimulated Peroxidase and Thiol Oxidase Activities: Pulse EPR and NMR Studies

Binding of Histidine in the (Cys)₃(His)₁-Coordinated [2Fe−2S] Cluster of Human mitoNEET

Comparison of EPR-Visible Cu2+ Sites in pMMO from Methylococcus capsulatus (Bath) and Methylomicrobium album BG8

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X‐band ESEEM spectroscopy of 15N substituted native and inhibitor‐bound superoxide dismutase

Cited by 10 publications

References 20 publications

Oxidation of Histidine Residues in Copper−Zinc Superoxide Dismutase by Bicarbonate-Stimulated Peroxidase and Thiol Oxidase Activities: Pulse EPR and NMR Studies

Oxidation of Histidine Residues in Copper−Zinc Superoxide Dismutase by Bicarbonate-Stimulated Peroxidase and Thiol Oxidase Activities: Pulse EPR and NMR Studies

Binding of Histidine in the (Cys)3(His)1-Coordinated [2Fe−2S] Cluster of Human mitoNEET

Comparison of EPR-Visible Cu2+ Sites in pMMO from Methylococcus capsulatus (Bath) and Methylomicrobium album BG8

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X‐band ESEEM spectroscopy of ¹⁵N substituted native and inhibitor‐bound superoxide dismutase

Binding of Histidine in the (Cys)₃(His)₁-Coordinated [2Fe−2S] Cluster of Human mitoNEET