STRUCTURE OF THE TOPA-SEMIQUINONE CATALYTIC INTERMEDIATE OF AMINE OXIDASE AS REVEALED BY MAGNETIC-INTERACTIONS WITH EXCHANGEABLE H-2 AND H-1 NUCLEI

被引:35
作者
WARNCKE, K
BABCOCK, GT
DOOLEY, DM
MCGUIRL, MA
MCCRACKEN, J
机构
[1] MICHIGAN STATE UNIV,DEPT CHEM,E LANSING,MI 48824
[2] MONTANA STATE UNIV,DEPT CHEM & BIOCHEM,BOZEMAN,MT 59717
关键词
D O I
10.1021/ja00088a042
中图分类号
O6 [化学];
学科分类号
0703 ;
摘要
Electron spin echo envelope modulation (ESEEM) and continuous wave-electron nuclear double resonance (CW-ENDOR) spectroscopies have been used selectively to characterize hyperfine interactions of aqueous solvent-exchangeable H-1 and H-2 nuclei with unpaired electron spin density in the topa-semiquinone catalytic intermediate of bovine amine oxidase. The radical was generated by anaerobic reduction of enzyme in (H2O)-H-2 or (H2O)-H-1 buffer with substrate benzylamine in the presence of cyanide ion. H-2 ESEEM spectra display a strong anisotropic hyperfine coupling of rhombic symmetry that is assigned to a single, exchangeable H-2 nucleus bonded alpha to the substrate-derived nitrogen atom that is incorporated into the cofactor during catalysis. Solvent exchange of this hydrogen dramatically influences the CW-electron paramagnetic resonance spectrum. ESEEM spectra show no evidence for deuteroxyl deuteron hyperfine coupling, suggesting that the two semiquinone oxygen atoms are deprotonated. Seven pairs of (H2O)-H-2 exchange-sensitive hyperfine couplings are observed in the H-1 CW-ENDOR spectrum. These couplings are assigned to protons involved in hydrogen bonds to the nitrogen and the two oxygen atoms of the radical. The exchange insensitivity of the H-1 CW-ENDOR matrix line and relatively weak H-2 matrix ESEEM demonstrate that the radical is well-sequestered from solvent water. The presence of a single proton covalently bonded to a trigonal nitrogen atom, and the p(p)i-orbital overlap in the C-N bond, shows that the nuclear and electronic structure of the active carbon center in the topa-semiquinone resembles that of the subsequent iminoquinone intermediate.
引用
收藏
页码:4028 / 4037
页数:10
相关论文
共 92 条
[21]   RECOVERY OF BROAD HYPERFINE LINES IN ELECTRON SPIN-ECHO ENVELOPE MODULATION SPECTROSCOPY OF DISORDERED-SYSTEMS [J].
FAUTH, JM ;
SCHWEIGER, A ;
ERNST, RR .
JOURNAL OF MAGNETIC RESONANCE, 1989, 81 (02) :262-274
[22]  
FERSHT A, 1985, ENZYME STRUCTURE MEC
[23]  
FINAZZOAGRO A, 1984, FEBS LETT, V214, P378
[24]   ELECTRON-NUCLEAR DOUBLE-RESONANCE STUDY OF FLAVODOXIN FROM PEPTOSTREPTOCOCCUS-ELSDENII [J].
FRITZ, J ;
MULLER, F ;
MAYHEW, SG .
HELVETICA CHIMICA ACTA, 1973, 56 (07) :2250-2254
[25]   SOLVENT EFFECTS IN ELECTRON SPIN RESONANCE SPECTRA [J].
GENDELL, J ;
FRAENKEL, GK ;
FREED, JH .
JOURNAL OF CHEMICAL PHYSICS, 1962, 37 (12) :2832-&
[26]  
Gordy W., 1980, TECHNIQUES CHEM, V15
[27]   STRUCTURE-FUNCTION STUDIES OF SUBSTRATE OXIDATION BY BOVINE SERUM AMINE OXIDASE - RELATIONSHIP TO COFACTOR STRUCTURE AND MECHANISM [J].
HARTMANN, C ;
KLINMAN, JP .
BIOCHEMISTRY, 1991, 30 (18) :4605-4611
[28]   SPECTROSCOPIC DETECTION OF CHEMICAL INTERMEDIATES IN THE REACTION OF PARA-SUBSTITUTED BENZYLAMINES WITH BOVINE SERUM AMINE OXIDASE [J].
HARTMANN, C ;
BRZOVIC, P ;
KLINMAN, JP .
BIOCHEMISTRY, 1993, 32 (09) :2234-2241
[29]  
HARTMANN C, 1987, J BIOL CHEM, V262, P962
[30]   PROTEIN TYROSYL RADICAL INTERACTIONS IN PHOTOSYSTEM-II STUDIED BY ELECTRON-SPIN-RESONANCE AND ELECTRON NUCLEAR DOUBLE-RESONANCE SPECTROSCOPY - COMPARISON WITH RIBONUCLEOTIDE REDUCTASE AND INVITRO TYROSINE [J].
HOGANSON, CW ;
BABCOCK, GT .
BIOCHEMISTRY, 1992, 31 (47) :11874-11880