title: Effects of protein structure on iron-polypeptide vibrational dynamic coupling in cytochrome c.
abstract: Cytochrome c (Cyt c) has a heme covalently bound to the polypeptide via a Cys-X-X-Cys-His (CXXCH) linker that is located in the interface region for protein-protein interactions. To determine whether the polypeptide matrix influences iron vibrational dynamics, nuclear resonance vibrational spectroscopy (NRVS) measurements were performed on (57)Fe-labeled ferric Hydrogenobacter thermophilus cytochrome c-552, and variants M13V, M13V/K22M, and A7F, which have structural modifications that alter the composition or environment of the CXXCH pentapeptide loop. Simulations of the NRVS data indicate that the 150-325 cm(-1) region is dominated by NHis-Fe-SMet axial ligand and polypeptide motions, while the 325-400 cm(-1) region shows dominant contributions from ν(Fe-NPyr) (Pyr = pyrrole) and other heme-based modes. Diagnostic spectral signatures that directly relate to structural features of the heme active site are identified using a quantum chemistry-centered normal coordinate analysis (QCC-NCA). In particular, spectral features that directly correlate with CXXCH loop stiffness, the strength of the Fe-His interaction, and the degree of heme distortion are identified. Cumulative results from our investigation suggest that compared to the wild type (wt), variants M13V and M13V/K22M have a more rigid CXXCH pentapeptide segment, a stronger Fe-NHis interaction, and a more ruffled heme. Conversely, the A7F variant has a more planar heme and a weaker Fe-NHis bond. These results are correlated to the observed changes in reduction potential between wt protein and the variants studied here. Implications of these results for Cyt c biogenesis and electron transfer are also discussed.
authors.affiliation: Department of Chemistry, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Mary Grace I
authors.initials: MG
authors.lastname: Galinato
authors.forename: Sarah E J
authors.initials: SE
authors.lastname: Bowman
authors.forename: Jesse G
authors.initials: JG
authors.lastname: Kleingardner
authors.forename: Sherri
authors.initials: S
authors.lastname: Martin
authors.forename: Jiyong
authors.initials: J
authors.lastname: Zhao
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Sturhahn
authors.forename: E Ercan
authors.initials: EE
authors.lastname: Alp
authors.forename: Kara L
authors.initials: KL
authors.lastname: Bren
authors.forename: Nicolai
authors.initials: N
authors.lastname: Lehnert
mesh_terms: D001665:Binding Sites / Q000502:physiology; D045304:Cytochromes c / Q000737:chemistry* / Q000378:metabolism*; D006454:Hemoglobins / Q000737:chemistry* / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism*; D017433:Protein Structure, Secondary; D014732:Vibration
chemical_list: D006454:Hemoglobins; D010455:Peptides; C570641:polypeptide-Fe; D045304:Cytochromes c