Spectroscopic comparisons of the pH dependencies of Fe-substituted (Mn)superoxide dismutase and Fe-superoxide dismutase.
Spectroscopic comparisons of the pH dependencies of Fe-substituted (Mn)superoxide dismutase and Fe-superoxide dismutase.
复制标题
Fe 取代 (Mn) 超氧化物歧化酶和 Fe 超氧化物歧化酶的 pH 依赖性的光谱比较。
DOI：
10.1021/bi972580r
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发表时间：
1998
期刊：
影响因子：
--
通讯作者：
Miller,AF
中科院分区：
文献类型：
--
作者：
Vance,CK;Miller,AF
We have compared the active sites ofEscherichia coliFe-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 pKis 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-, theKDfor 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-inThermusthermophilusMn-SOD than Fe-SOD [Lah, M., et al. (1995)Biochemistry 34, 1646−1660]. We find a pKof 8.5 to be associated with dramatic changes in the EPR spectrum. In addition, we confirm the pKbetween 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 pKappears 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 lowE° is due to perturbation of the oxidized state.
