title: G-tensors of the flavin adenine dinucleotide radicals in glucose oxidase: a comparative multifrequency electron paramagnetic resonance and electron-nuclear double resonance study.
abstract: 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) at high microwave frequencies (93.9 and 360 GHz) and correspondingly high magnetic fields and by pulsed electron-nuclear double resonance (ENDOR) spectroscopy at 9.7 GHz. Because of the high spectral resolution of the frozen-solution continuous-wave EPR spectrum recorded at 360 GHz, the anisotropy of the g-tensor of FAD*- could be fully resolved. By least-squares fittings of spectral simulations to experimental data, the principal 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 more axially symmetric as compared to that of FADH*. In contrast, significantly altered proton hyperfine couplings were observed by ENDOR upon transforming the neutral FADH* radical into the anionic FAD*- radical by pH titration of GO. That the g-principal values of both protonation forms remain largely identical demonstrates the 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 ideally suited to identify chemically different flavin radicals.
authors.affiliation: Freie Universität Berlin, Fachbereich Physik, Institut für Experimentalphysik, Arnimallee 14, 14195 Berlin, Germany.
authors.forename: Asako
authors.initials: A
authors.lastname: Okafuji
authors.forename: Alexander
authors.initials: A
authors.lastname: Schnegg
authors.forename: Erik
authors.initials: E
authors.lastname: Schleicher
authors.forename: Klaus
authors.initials: K
authors.lastname: Möbius
authors.forename: Stefan
authors.initials: S
authors.lastname: Weber
mesh_terms: D000465:Algorithms; D016880:Anisotropy; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods; D005182:Flavin-Adenine Dinucleotide / Q000737:chemistry* / Q000378:metabolism; D005609:Free Radicals / Q000737:chemistry* / Q000378:metabolism; D005949:Glucose Oxidase / Q000737:chemistry* / Q000378:metabolism; D016018:Least-Squares Analysis; D015394:Molecular Structure; D011522:Protons; D017186:Titrimetry / Q000379:methods
chemical_list: D005609:Free Radicals; D011522:Protons; D005182:Flavin-Adenine Dinucleotide; D005949:Glucose Oxidase