EPR, ENDOR, and Special TRIPLE measurements of P•+ in wild type and modified reaction centers from Rb. sphaeroides

J. P. Allen, J. M. Cordova, C. C. Jolley, T. A. Murray, J. W. Schneider, N. W. Woodbury, J. C. Williams, J. Niklas, G. Klihm, M. Reus, W. Lubitz

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Abstract

The influence of the protein environment on the primary electron donor, P, a bacteriochlorophyll a dimer, of reaction centers from Rhodobacter sphaeroides, has been investigated using electron paramagnetic resonance and electron nuclear double resonance spectroscopy. These techniques were used to probe the effects on P that are due to alteration of three amino acid residues, His L168, Asn L170, and Asn M199. The introduction of Glu at L168, Asp at L170, or Asp at M199 changes the oxidation/reduction midpoint potential of P in a pH-dependent manner (Williams et al. (2001) Biochemistry 40, 15403-15407). For the double mutant His L168 to Glu and Asn at L170 to Asp, excitation results in electron transfer along the A-side branch of cofactors at pH 7.2, but at pH 9.5, a long-lived state involving B-side cofactors is produced (Haffa et al. (2004) J Phys Chem B 108, 4-7). Using electron paramagnetic resonance spectroscopy, the mutants with alterations of each of the three individual residues and a double mutant, with changes at L168 and L170, were found to have increased linewidths of 10.1-11.0 G compared to the linewidth of 9.6 G for wild type. The Special TRIPLE spectra were pH dependent, and at pH 8, the introduction of aspartate at L170 increased the spin density ratio, ρ LM, to 6.1 while an aspartate at the symmetry related position, M199, decreased the ratio to 0.7 compared to the value of 2.1 for wild type. These results indicate that the energy of the two halves of P changes by about 100 meV due to the mutations and are consistent with the interpretation that electrostatic interactions involving these amino acid residues contribute to the switch in pathway of electron transfer.

Original languageEnglish
Pages (from-to)1-10
Number of pages10
JournalPhotosynthesis Research
Volume99
Issue number1
DOIs
Publication statusPublished - Jan 2009

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Electron Spin Resonance Spectroscopy
Paramagnetic resonance
electron paramagnetic resonance spectroscopy
Electrons
Aspartic Acid
Linewidth
aspartic acid
Viperidae
mutants
electron transfer
Spectroscopy
Bacteriochlorophylls
Spectrum Analysis
Amino Acids
Biochemistry
Rhodobacter sphaeroides
Coulomb interactions
amino acids
Dimers
electrostatic interactions

Keywords

  • Bacteriochlorophyll
  • Electron paramagnetic resonance
  • Magnetic resonance
  • Oxidized bacteriochlorophyll dimer
  • Purple bacteria
  • Reaction centers

ASJC Scopus subject areas

  • Plant Science
  • Cell Biology
  • Biochemistry

Cite this

EPR, ENDOR, and Special TRIPLE measurements of P•+ in wild type and modified reaction centers from Rb. sphaeroides. / Allen, J. P.; Cordova, J. M.; Jolley, C. C.; Murray, T. A.; Schneider, J. W.; Woodbury, N. W.; Williams, J. C.; Niklas, J.; Klihm, G.; Reus, M.; Lubitz, W.

In: Photosynthesis Research, Vol. 99, No. 1, 01.2009, p. 1-10.

Research output: Contribution to journalArticle

Allen, JP, Cordova, JM, Jolley, CC, Murray, TA, Schneider, JW, Woodbury, NW, Williams, JC, Niklas, J, Klihm, G, Reus, M & Lubitz, W 2009, 'EPR, ENDOR, and Special TRIPLE measurements of P•+ in wild type and modified reaction centers from Rb. sphaeroides', Photosynthesis Research, vol. 99, no. 1, pp. 1-10. https://doi.org/10.1007/s11120-008-9346-6
Allen, J. P. ; Cordova, J. M. ; Jolley, C. C. ; Murray, T. A. ; Schneider, J. W. ; Woodbury, N. W. ; Williams, J. C. ; Niklas, J. ; Klihm, G. ; Reus, M. ; Lubitz, W. / EPR, ENDOR, and Special TRIPLE measurements of P•+ in wild type and modified reaction centers from Rb. sphaeroides. In: Photosynthesis Research. 2009 ; Vol. 99, No. 1. pp. 1-10.
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abstract = "The influence of the protein environment on the primary electron donor, P, a bacteriochlorophyll a dimer, of reaction centers from Rhodobacter sphaeroides, has been investigated using electron paramagnetic resonance and electron nuclear double resonance spectroscopy. These techniques were used to probe the effects on P that are due to alteration of three amino acid residues, His L168, Asn L170, and Asn M199. The introduction of Glu at L168, Asp at L170, or Asp at M199 changes the oxidation/reduction midpoint potential of P in a pH-dependent manner (Williams et al. (2001) Biochemistry 40, 15403-15407). For the double mutant His L168 to Glu and Asn at L170 to Asp, excitation results in electron transfer along the A-side branch of cofactors at pH 7.2, but at pH 9.5, a long-lived state involving B-side cofactors is produced (Haffa et al. (2004) J Phys Chem B 108, 4-7). Using electron paramagnetic resonance spectroscopy, the mutants with alterations of each of the three individual residues and a double mutant, with changes at L168 and L170, were found to have increased linewidths of 10.1-11.0 G compared to the linewidth of 9.6 G for wild type. The Special TRIPLE spectra were pH dependent, and at pH 8, the introduction of aspartate at L170 increased the spin density ratio, ρ L/ρ M, to 6.1 while an aspartate at the symmetry related position, M199, decreased the ratio to 0.7 compared to the value of 2.1 for wild type. These results indicate that the energy of the two halves of P changes by about 100 meV due to the mutations and are consistent with the interpretation that electrostatic interactions involving these amino acid residues contribute to the switch in pathway of electron transfer.",
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AU - Schneider, J. W.

AU - Woodbury, N. W.

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