Research Article

Non-planar heme deformations and excited state displacements in horseradish peroxidase detected by Raman spectroscopy at Soret excitation

  1. Qing Huang1,2,
  2. Reinhard Schweitzer-Stenner1,*

Article first published online: 15 APR 2005

DOI: 10.1002/jrs.1326

Issue

Journal of Raman Spectroscopy

Journal of Raman Spectroscopy

Special Issue: Structure and dynamics of biomolecules

Volume 36, Issue 4, pages 363–375, April 2005

Additional Information

How to Cite

Huang, Q. and Schweitzer-Stenner, R. (2005), Non-planar heme deformations and excited state displacements in horseradish peroxidase detected by Raman spectroscopy at Soret excitation. J. Raman Spectrosc., 36: 363–375. doi: 10.1002/jrs.1326

Author Information

  1. 1

    Department of Chemistry, Drexel University, 3141 Chestnut Street, Philadelphia, Pennsylvania 19104, USA

  2. 2

    Department of Chemistry, University of Puerto Rico, Río Piedras Campus, San Juan, Puerto Rico 00931, USA

*Department of Chemistry, Drexel University, 3141 Chestnut Street, Philadelphia, Pennsylvania 19104, USA.

Publication History

  1. Issue published online: 15 APR 2005
  2. Article first published online: 15 APR 2005
  3. Manuscript Accepted: 2 FEB 2005
  4. Manuscript Received: 3 DEC 2004

Funded by

  • National Science Foundation. Grant Number: MCB-0318749.
  • ACS-Petroleum Research Fund. Grant Number: PRF 37406-AC.
  • NIH. Grant Number: P20 RR16439-01.

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Keywords:

  • vibronic coupling;
  • Franck–Condon interaction;
  • heme protein;
  • non-planar distortions;
  • absorption;
  • resonance Raman spectroscopy

Abstract

We studied the out-of-plane modes of horseradish peroxidase (HRP) in different spin, oxidation and ligation states by measuring the respective resonance Raman spectra with (near) Soret excitation. The non-planar modes of the heme are Raman inactive for planar macrocycles, but become resonance Raman active in the presence of out-of-plane deformations. The thus induced Raman scattering results mostly from Franck–Condon type coupling. We observed bands from a variety of out-of-plane modes such as γ5, γ6, γ7(A2u) modes, γ21 and γ22(Eg), γ15(B2u) and γ11(B1u). The appearance of these bands is clearly indicative of non-planar deformations of the same symmetry. We determined the relative intensities of all sufficiently intense Raman modes of ferric pentacoordinated, quantum mixed and hexacoordinated low-spin as well as of ferrous pentacoordinated high-spin HRP C. The apparent vibronic coupling parameters were obtained from a self-consistent analysis of the Raman intensities and the respective optical absorption spectrum. They revealed significantly reduced displacements of the excited B-state for the ferrous, deoxy-like state compared with the resting ferric state. An analysis of the obtained coupling strengths of out-of-plane modes revealed that they are predominantly induced by static non-planar deformations along the normal coordinates of the lowest wavenumber modes of a given symmetry type. We used the deformations of the heme in resting HRP C to determine the vibronic coupling strength of the experimentally detectable out-of-plane modes and subsequently employed this information to obtain the non-planar deformations for the investigated ferric hexacoordinated low-spin state. This yielded a substantial reduction of the ruffling distortion, whereas the doming deformation remained mostly unaffected by the change of the iron's spin and ligation state. Copyright © 2005 John Wiley & Sons, Ltd.

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