G-Tensors of the Flavin Adenine Dinucleotide Radicals in Glucose Oxidase: A Comparative Multifrequency Electron Paramagnetic Resonance and Electron-Nuclear Double Resonance Study
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- 作者：Asako Okafuji ; Alexander Schnegg ; Erik Schleicher ; Klaus Mö ; bius ; Stefan Weber
- 刊名：Journal of Physical Chemistry B
- 出版年：2008
- 出版时间：March 20, 2008
- 年：2008
- 卷：112
- 期：11
- 页码：3568 - 3574
- 全文大小：115K
- 年卷期：v.112,no.11(March 20, 2008)
- ISSN：1520-5207
文摘
The flavin adenine dinucleotide (FAD) cofactor of Aspergillus niger glucose oxidase (GO) in its anionic(FAD
-) and neutral (FADH) radical form was investigated by electron paramagnetic resonance (EPR) athigh microwave frequencies (93.9 and 360 GHz) and correspondingly high magnetic fields and by pulsedelectron-nuclear double resonance (ENDOR) spectroscopy at 9.7 GHz. Because of the high spectral resolutionof the frozen-solution continuous-wave EPR spectrum recorded at 360 GHz, the anisotropy of the g-tensor ofFAD- could be fully resolved. By least-squares fittings of spectral simulations to experimental data, theprincipal values of g have been established with high precision: gX = 2.00429(3), gY = 2.00389(3), gZ =2.00216(3) (X, Y, and Z are the principal axes of g) yielding giso = 2.00345(3). The gY-component of FAD-from GO is moderately shifted upon deprotonation of FADH, rendering the g-tensor of FAD- slightly moreaxially symmetric as compared to that of FADH. In contrast, significantly altered proton hyperfine couplingswere observed by ENDOR upon transforming the neutral FADH radical into the anionic FAD- radical bypH titration of GO. That the g-principal values of both protonation forms remain largely identical demonstratesthe robustness of g against local changes in the electron-spin density distribution of flavins. Thus, in flavins,the g-tensor reflects more global changes in the electronic structure and, therefore, appears to be ideallysuited to identify chemically different flavin radicals.
-) and neutral (FADH) radical form was investigated by electron paramagnetic resonance (EPR) athigh microwave frequencies (93.9 and 360 GHz) and correspondingly high magnetic fields and by pulsedelectron-nuclear double resonance (ENDOR) spectroscopy at 9.7 GHz. Because of the high spectral resolutionof the frozen-solution continuous-wave EPR spectrum recorded at 360 GHz, the anisotropy of the g-tensor ofFAD- could be fully resolved. By least-squares fittings of spectral simulations to experimental data, theprincipal values of g have been established with high precision: gX = 2.00429(3), gY = 2.00389(3), gZ =2.00216(3) (X, Y, and Z are the principal axes of g) yielding giso = 2.00345(3). The gY-component of FAD-from GO is moderately shifted upon deprotonation of FADH, rendering the g-tensor of FAD- slightly moreaxially symmetric as compared to that of FADH. In contrast, significantly altered proton hyperfine couplingswere observed by ENDOR upon transforming the neutral FADH radical into the anionic FAD- radical bypH titration of GO. That the g-principal values of both protonation forms remain largely identical demonstratesthe robustness of g against local changes in the electron-spin density distribution of flavins. Thus, in flavins,the g-tensor reflects more global changes in the electronic structure and, therefore, appears to be ideallysuited to identify chemically different flavin radicals.