The Solution Structure of Oxidized HiPIP I from Ectothiorhodospira halophila; Can NMR Spectroscopy Be Used to Probe Rearrangements Associated with Electron Transfer Processes?

Abstract

AbstractIn the 1H NMR spectrum of the oxidized form of the high‐potential iron‐sulfur protein (HiPIPI) from Ectothiorhodospira halophila, 91% of the total proton resonances and 100% of the residues have been assigned. The standard COSY, NOESY, and TOCSY sequences have been optimized for the paramagnetism of the molecule. Extensive assignment of the 15N NMR spectrum has been obtained through HMQC spectra. With 1437 dipolar connectivities, of which about 10% involved fast‐relaxing protons, a family of 18 structures was generated with an RMSD of 0.65 Å by using the programs developed by Wüthrich. The family of structures was further refined by various calculation steps; the final RMSD was 0.48 Å. The structures appear to be very similar but not equal to the structures of the reduced protein. Despite the similarity in structure, significant variations in the chemical shifts are observed. A similar behavior was observed for the homologous protein from Chromatium vinosum. It is concluded that NMR is a sensitive tool to monitor differences between oxidized and reduced proteins; however, the detailed structural variations should be evaluated with caution at the present level of resolution, which roughly corresponds to a resolution of 2.5 Å in an X‐ray structure.