Oxo-vanadium as a spin probe for the investigation of the metal coordination environment of imidazole glycerol phosphate dehydratase

Imidazole glycerol phosphate dehydratase (IGPD) catalyses the dehydration of imidazole glycerol phosphate to imidazole acetol phosphate, an important late step in the biosynthesis of histidine. IGPD, isolated as a low molecular weight and inactive ape-form, assembles with specific divalent metal cations to form a catalytically active high molecular weight metalloenzyme. Oxo-vanadium ions also assemble the protein into, apparently, the same high molecular weight form but, uniquely, yield a protein without catalytic activity. The VO2+ derivative of IGPD has been investigated by electron paramagnetic resonance (EPR), electron nuclear double resonance (ENDOR) and electron spin echo envelope modulation (ESEEM) spectroscopy. The spin Hamiltonian parameters indicate the presence of multiple N-14 nuclei in the inner coordination sphere of VO2+ which is corroborated by ENDOR and ESEEM spectra showing resonances attributable to interactions with N-14 nuclei. The isotropic superhyperfine coupling component of about 7 MHz determined by ENDOR is consistent with a nitrogen of coordinated histidine imidazole(s). The ESEEM fourier-transform spectra further support the notion that the VO2+ substituted enzyme contains inner-sphere nitrogen ligands. The isotropic and anisotropic N-14 superhyperfine coupling components are similar to those reported for other equatorially coordinated enzymatic histidine imidazole systems. ESEEM resonances from axial N-14 ligands are discussed. (C) 2000 Elsevier Science Inc. All rights reserved.