Spectroscopic comparisons of the pH dependencies of Fe-substituted (Mn) superoxide dismutase and Fe-superoxide dismutase
Abstract
We have compared the active sites of Escherichia coli Fe-substituted (Mn)superoxide dismutase [Fe-sub-(Mn)SOD] and Fe-SOD to elucidate the basis for the inactivity of Fe-sub-(Mn)SOD, despite its apparent similarity to Fe-SOD. The active site of (reduced) Fe2+-sub-(Mn)SOD is qualitatively similar to that of native Fe2+-SOD, indicating similar active site structures and coordination environments for Fe2+. Its nativelike pK is indicative of nativelike local electrostatics, and consistent with Fe2+-sub-(Mn)SOD's retention of ability to reduce O2*- [Vance and Miller (1998) J. Am. Chem. Soc. 120(3), 461-467]. The active site of (oxidized) Fe3+-sub-(Mn)SOD differs from that of Fe3+-SOD with respect to the EPR signals produced at both neutral and high pH, indicating different coordination environments for Fe3+. Although Fe3+-sub-(Mn)SOD binds the small anions N3- and F-, the KD for N3- is tighter than that of Fe3+-SOD, suggesting that the (Mn)SOD protein favors anion binding more than does the (Fe)SOD protein. The EPR spectral consequences of binding F- are reminiscent of those observed upon binding the first F- to Fe3+-SOD, but the EPR spectrum obtained upon binding N3- is different, consistent with crystallographic observation of a different binding mode for N3- in Thermus thermophilus Mn-SOD than Fe-SOD [Lah, M., et al. (1995) Biochemistry 34, 1646-1660]. We find a pK of 8.5 to be associated with dramatic changes in the EPR spectrum. In addition, we confirm the pK between 6 and 7 that has previously been reported based on changes in the optical signal and N3- binding [Yamakura, F., et al. (1995) Eur. J. Biochem. 227, 700-706]. However, this latter pK appears to be associated with much subtler changes in the EPR spectrum. The non-native pKs observed in Fe3+-sub-(Mn)SOD and the differences in the Fe3+ coordination indicated by the EPR spectra are consistent with Fe3+-sub-(Mn)SOD's inability to oxidize O2*- and suggest that its low E degrees is due to perturbation of the oxidized state.
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Using pulse radiolysis studies, the following mechanism has been proposed.
“Using pulse radiolysis studies, the following mechanism has been proposed [84,85], Figure 36.”
From Superoxide Anion Chemistry—Its Role at the Core of the Innate Immunity (2023) · cites this paper as [85]
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- Authors
- Vance, C. K., & Miller, A.-F.
- Affiliations
- Johns Hopkins University (US)
- Corresponding author
- Miller, Anne-Frances
- Venue
- Biochemistry
- Publisher
- American Chemical Society (ACS)
- Published
- 1998-04-01
- Volume
- 37
- Issue
- 16
- Pages
- 5518-5527
- ISSN
- 0006-2960 (ISSN-L)
- 1520-4995
- OpenAlex
- W1977875170
- Cited by
- 89
- References
- 17
- Citations by year
- 2026: 1 · 2025: 1 · 2024: 1 · 2023: 2 · 2022: 1 · 2021: 1 · 2019: 1 · 2018: 2 · 2017: 7 · 2016: 2 · 2015: 2 · 2014: 6 · 2013: 5 · 2012: 2
- FWCI
- 3.51
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Biochemistry (publishedVersion)
- PubMed (publishedVersion)
- Funders
- National Institutes of Health (1s10rr004794-01)
- National Institute of General Medical Sciences (T32-GM07231, T32GM007231)
- National Center for Research Resources (RR 04794)
- Topics
- Physical Sciences › Chemistry › Inorganic Chemistry › Metal-Catalyzed Oxygenation Mechanisms
- Physical Sciences › Energy › Renewable Energy, Sustainability and the Environment › Metalloenzymes and iron-sulfur proteins
- Physical Sciences › Materials Science › Materials Chemistry › Enzyme Structure and Function
- Keywords
- superoxide dismutase, manganese superoxide dismutase, iron superoxide dismutase, pH dependence, active site, iron coordination environment, electron spin resonance, nitrate binding, fluoride binding, anion binding, pK, Escherichia coli, Thermus thermophilus
- MeSH terms
- Binding Sites, Electron Spin Resonance Spectroscopy, Ferrous Compounds, Hydrogen-Ion Concentration, Iron, Kinetics, Manganese, Oxidation-Reduction, Substrate Specificity, Superoxide Dismutase, Titrimetry
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