Paramagnetic properties of the low- and high-spin states of yeast cytochrome c peroxidase
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Here we describe paramagnetic NMR analysis of the low- and high-spin forms of yeast cytochrome c peroxidase (CcP), a 34 kDa heme enzyme involved in hydroperoxide reduction in mitochondria. Starting from the assigned NMR spectra of a low-spin CN-bound CcP and using a strategy based on paramagnetic pseudocontact shifts, we have obtained backbone resonance assignments for the diamagnetic, iron-free protein and the high-spin, resting-state enzyme. The derived chemical shifts were further used to determine low- and high-spin magnetic susceptibility tensors and the zero-field splitting constant (D) for the high-spin CcP. The D value indicates that the latter contains a hexacoordinate heme species with a weak field ligand, such as water, in the axial position. Being one of the very few high-spin heme proteins analyzed in this fashion, the resting state CcP expands our knowledge of the heme coordination chemistry in biological systems.
KeywordsParamagnetic NMR Heme proteins Pseudocontact shifts Zero-field splitting
A.N.V. is an FWO Post-Doctoral Researcher. We thank Yann Streckx for the critical reading of the manuscript. This work was supported by the VIB and the Flemish Hercules Foundation.
- Bosshard HR, Anni H, Yonetani T (1990) Yeast cytochrome c peroxidase. In: Everse J, Everse KE, Grisham MB (eds) Peroxidases in chemistry and biology. CRC Press, Boca Raton, pp 51–84Google Scholar
- Edwards SL, Poulos TL (1990) Ligand binding and structural perturbations in cytochrome c peroxidase. A crystallographic study. J Biol Chem 265:2588–2595Google Scholar
- La Mar GN, Satterlee J, de Ropp JS (2000) Nuclear magnetic resonance of hemoproteins. In: Kadish KM, Smith KM, Guilard R (eds) The porphyrin handbook. Academic press, San Diego, pp 185–298Google Scholar
- Walker FA (2000) Proton NMR and EPR spectroscopy of paramagnetic metalloporphyrins. In: Kadish KM, Smith KM, Guilard R (eds) The porphyrin handbook. Academic press, San Diego, pp 81–184Google Scholar
- Yonetani T, Anni H (1987) Yeast cytochrome c peroxidase. Coordination and spin states of heme prosthetic group. J Biol Chem 262:9547–9554Google Scholar