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.
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