Identification of Dimethylbenzimidazole Axial Coordination and Characterization of 14N Superhyperfine and Nuclear Quadrupole Coupling in Cob(II)alamin Bound to Ethanolamine Deaminase in a C
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- 作者：Shyue-Chu Ke ; Maricel Torrent ; Djamaladdin G. Museav ; Keiji Morokuma ; and Kurt Warncke
- 刊名：Biochemistry
- 出版年：1999
- 出版时间：September 28, 1999
- 年：1999
- 卷：38
- 期：39
- 页码：12681 - 12689
- 全文大小：102K
- 年卷期：v.38,no.39(September 28, 1999)
- ISSN：1520-4995
文摘
Cobalt(II)-14N superhyperfine and 14N nuclear quadrupole couplings in cryotrapped free andethanolamine deaminase-bound cob(II)alamin have been characterized in the disordered solid state byusing X-band electron spin-echo envelope modulation (ESEEM) spectroscopy. Enzyme-bound cob(II)alamin was cryotrapped after formation by substrate-initiated, thermally activated cleavage of the cobalt-carbon bond of adenosylcobalamin. Free dimethylbenzimidazole axial base-on cob(II)alamin was formedby photolysis of the corresponding adenosylcobalamin and cryotrapped in glycerol-aqueous glass. Three-pulse ESEEM experiments were performed by using microwave pulse excitation at the g
value of CoIIat magnetic field values of 287.0 and 345.0 mT and over a range of 
values from 227 to 1316 ns. Twocommon sets of 14N features are distinguished in the ESEEM spectra. One set is assigned to the remote(N1) nitrogen in the dimethylbenzimidazole 
-axial ligand by using two independent approaches: (a)comparison of ESEEM from cob(II)alamin with ESEEM from cob(II)inamide-ligand model compoundsand (b) from the correspondence between the N1 14N nuclear quadrupole parameters derived from ESEEMsimulations and those computed by using density functional theory. The second set is assigned to thecorrin ring 14N nuclei. The results identify the coenzyme's on-board dimethylbenzimidazole moiety asthe -axial ligand to cob(II)alamin in ethanolamine deaminase in the substrate radical-CoII biradicalcatalytic intermediate state. Thus, CoII is a pentacoordinate, -axial liganded complex during turnover.We infer that dimethylbenzimidazole is also the -axial ligand to the intact coenzyme in the restingenzyme. A 14% increase in the isotropic hyperfine coupling of the remote dimethylbenzimidazole 14Nnucleus in enzyme-bound versus free base-on cob(II)alamin shows an enhanced delocalization of unpairedspin density from CoII onto the axial ligand, which would contribute to the acceleration of the cobalt-carbon bond cleavage rate in situ.
value of CoIIat magnetic field values of 287.0 and 345.0 mT and over a range of values from 227 to 1316 ns. Twocommon sets of 14N features are distinguished in the ESEEM spectra. One set is assigned to the remote(N1) nitrogen in the dimethylbenzimidazole -axial ligand by using two independent approaches: (a)comparison of ESEEM from cob(II)alamin with ESEEM from cob(II)inamide-ligand model compoundsand (b) from the correspondence between the N1 14N nuclear quadrupole parameters derived from ESEEMsimulations and those computed by using density functional theory. The second set is assigned to thecorrin ring 14N nuclei. The results identify the coenzyme's on-board dimethylbenzimidazole moiety asthe -axial ligand to cob(II)alamin in ethanolamine deaminase in the substrate radical-CoII biradicalcatalytic intermediate state. Thus, CoII is a pentacoordinate, -axial liganded complex during turnover.We infer that dimethylbenzimidazole is also the -axial ligand to the intact coenzyme in the restingenzyme. A 14% increase in the isotropic hyperfine coupling of the remote dimethylbenzimidazole 14Nnucleus in enzyme-bound versus free base-on cob(II)alamin shows an enhanced delocalization of unpairedspin density from CoII onto the axial ligand, which would contribute to the acceleration of the cobalt-carbon bond cleavage rate in situ.