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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 57Fe-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 N His–Fe–S Met axial ligand and polypeptide motions, while the 325–400 cm –1 region shows dominant contributions from ν(Fe–N Pyr) (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–N His interaction, and a more ruffled heme. Conversely, themore » 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. Lastly, we discuss the implications of these results for Cyt c biogenesis and electron transfer.« less

Authors:
 [1];  [2];  [2];  [3];  [4];  [4];  [4];  [2];  [3]
  1. Univ. of Michigan, Ann Arbor, MI (United States). Dept. of Chemistry; Penn State, Erie, PA (United States). Behrend College
  2. Univ. of Rochester, NY (United States). Dept. of Chemistry
  3. Univ. of Michigan, Ann Arbor, MI (United States). Dept. of Chemistry
  4. Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States)
Sponsoring Org.:
National Institutes of Health (NIH); USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22)
OSTI Identifier:
1261133
Alternate Identifier(s):
OSTI ID: 1362102
Grant/Contract Number:  
R01-GM63170; AC02-06CH11357
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Biochemistry
Additional Journal Information:
Journal Volume: 54; Journal Issue: 4; Journal ID: ISSN 0006-2960
Publisher:
American Chemical Society (ACS)
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Galinato, Mary Grace I., Bowman, Sarah E. J., Kleingardner, Jesse G., Martin, Sherri, Zhao, Jiyong, Sturhahn, Wolfgang, Alp, E. Ercan, Bren, Kara L., and Lehnert, Nicolai. Effects of Protein Structure on Iron–Polypeptide Vibrational Dynamic Coupling in Cytochrome c. United States: N. p., 2014. Web. doi:10.1021/bi501430z.
Galinato, Mary Grace I., Bowman, Sarah E. J., Kleingardner, Jesse G., Martin, Sherri, Zhao, Jiyong, Sturhahn, Wolfgang, Alp, E. Ercan, Bren, Kara L., & Lehnert, Nicolai. Effects of Protein Structure on Iron–Polypeptide Vibrational Dynamic Coupling in Cytochrome c. United States. doi:10.1021/bi501430z.
Galinato, Mary Grace I., Bowman, Sarah E. J., Kleingardner, Jesse G., Martin, Sherri, Zhao, Jiyong, Sturhahn, Wolfgang, Alp, E. Ercan, Bren, Kara L., and Lehnert, Nicolai. Mon . "Effects of Protein Structure on Iron–Polypeptide Vibrational Dynamic Coupling in Cytochrome c". United States. doi:10.1021/bi501430z. https://www.osti.gov/servlets/purl/1261133.
@article{osti_1261133,
title = {Effects of Protein Structure on Iron–Polypeptide Vibrational Dynamic Coupling in Cytochrome c},
author = {Galinato, Mary Grace I. and Bowman, Sarah E. J. and Kleingardner, Jesse G. and Martin, Sherri and Zhao, Jiyong and Sturhahn, Wolfgang and Alp, E. Ercan and Bren, Kara L. and Lehnert, Nicolai},
abstractNote = {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 57Fe-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. Lastly, we discuss the implications of these results for Cyt c biogenesis and electron transfer.},
doi = {10.1021/bi501430z},
journal = {Biochemistry},
issn = {0006-2960},
number = 4,
volume = 54,
place = {United States},
year = {2014},
month = {12}
}

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