Abstract
Site-specific mutations in the quinone binding sites of the photosynthetic reaction center (RC) protein complexes of Rhodobacter (R.) capsulatus caused pronounced effects on sequential electron transfer. Conserved residues that break the twofold symmetry in this region of the RC - M246Ala and M247Ala in the Q(A) binding pocket, and L212Glu and L213Asp in the Q(B) binding pocket - were targeted. We constructed a Q(B)-site mutant, L212Glu-L213Asp → Ala-Ala, and a Q(A)-site mutant, M246Ala-M247Ala → Glu-Asp, to partially balance the differences in charge distribution normally found between the two quinone binding sites. In addition, two photocompetent revertants were isolated from the photosynthetically-incompetent M246Glu-M247Asp mutant: M246Ala-M247Asp and M246Gly-M247Asp. Sequential electron transfer was investigated by continuous light excitation and time-resolved electron paramagnetic resonance (EPR), and time-resolved optical techniques. Several lines of EPR evidence suggested that the forward electron transfer rate to Q(A), k(Q), was slowed in those strains containing altered Q(A) sites. The slower rates of secondary electron transfer were confirmed by time-resolved optical results with the M246Glu-M247Asp mutations in the Q(A) site resulting in a dramatically lowered secondary electron transfer efficiency [k(Q) <(2 ns)-1] in comparison with either the native R. capsulatus RC or the Q(B) site mutant [k(Q) ≃ (200 ps)-1]. Secondary electron transfer in the two revertants was intermediate between that of the native RC and the Q(A) mutant. The P+Q(A)- → PQ(A) charge recombination rates were also changed in the strains that carried altered Q(A) sites. We show that local mutations in the Q(A) site, presumably through local electrostatic changes, significantly alter binding and electron transfer properties of Q(A).
| Original language | English |
|---|---|
| Pages (from-to) | 93-103 |
| Number of pages | 11 |
| Journal | Photosynthesis Research |
| Volume | 52 |
| Issue number | 2 |
| DOIs | |
| Publication status | Published - 1997 |
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Keywords
- Electron paramagnetic resonance
- Electron spin polarization
- Electrostatics
- Quinone binding
- Revertant isolation
ASJC Scopus subject areas
- Plant Science
Cite this
Spectroscopic characterization of quinone-site mutants of the bacterial photosynthetic reaction center. / Laible, Philip D.; Zhang, Yuenian; Morris, Andrea L.; Snyder, Seth W.; Ainsworth, Clint; Greenfield, Scott R.; Wasielewski, Michael R; Parot, Pierre; Schoepp, Barbara; Schiffer, Marianne; Hanson, Deborah K.; Thurnauer, Marion C.
In: Photosynthesis Research, Vol. 52, No. 2, 1997, p. 93-103.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Spectroscopic characterization of quinone-site mutants of the bacterial photosynthetic reaction center
AU - Laible, Philip D.
AU - Zhang, Yuenian
AU - Morris, Andrea L.
AU - Snyder, Seth W.
AU - Ainsworth, Clint
AU - Greenfield, Scott R.
AU - Wasielewski, Michael R
AU - Parot, Pierre
AU - Schoepp, Barbara
AU - Schiffer, Marianne
AU - Hanson, Deborah K.
AU - Thurnauer, Marion C.
PY - 1997
Y1 - 1997
N2 - Site-specific mutations in the quinone binding sites of the photosynthetic reaction center (RC) protein complexes of Rhodobacter (R.) capsulatus caused pronounced effects on sequential electron transfer. Conserved residues that break the twofold symmetry in this region of the RC - M246Ala and M247Ala in the Q(A) binding pocket, and L212Glu and L213Asp in the Q(B) binding pocket - were targeted. We constructed a Q(B)-site mutant, L212Glu-L213Asp → Ala-Ala, and a Q(A)-site mutant, M246Ala-M247Ala → Glu-Asp, to partially balance the differences in charge distribution normally found between the two quinone binding sites. In addition, two photocompetent revertants were isolated from the photosynthetically-incompetent M246Glu-M247Asp mutant: M246Ala-M247Asp and M246Gly-M247Asp. Sequential electron transfer was investigated by continuous light excitation and time-resolved electron paramagnetic resonance (EPR), and time-resolved optical techniques. Several lines of EPR evidence suggested that the forward electron transfer rate to Q(A), k(Q), was slowed in those strains containing altered Q(A) sites. The slower rates of secondary electron transfer were confirmed by time-resolved optical results with the M246Glu-M247Asp mutations in the Q(A) site resulting in a dramatically lowered secondary electron transfer efficiency [k(Q) <(2 ns)-1] in comparison with either the native R. capsulatus RC or the Q(B) site mutant [k(Q) ≃ (200 ps)-1]. Secondary electron transfer in the two revertants was intermediate between that of the native RC and the Q(A) mutant. The P+Q(A)- → PQ(A) charge recombination rates were also changed in the strains that carried altered Q(A) sites. We show that local mutations in the Q(A) site, presumably through local electrostatic changes, significantly alter binding and electron transfer properties of Q(A).
AB - Site-specific mutations in the quinone binding sites of the photosynthetic reaction center (RC) protein complexes of Rhodobacter (R.) capsulatus caused pronounced effects on sequential electron transfer. Conserved residues that break the twofold symmetry in this region of the RC - M246Ala and M247Ala in the Q(A) binding pocket, and L212Glu and L213Asp in the Q(B) binding pocket - were targeted. We constructed a Q(B)-site mutant, L212Glu-L213Asp → Ala-Ala, and a Q(A)-site mutant, M246Ala-M247Ala → Glu-Asp, to partially balance the differences in charge distribution normally found between the two quinone binding sites. In addition, two photocompetent revertants were isolated from the photosynthetically-incompetent M246Glu-M247Asp mutant: M246Ala-M247Asp and M246Gly-M247Asp. Sequential electron transfer was investigated by continuous light excitation and time-resolved electron paramagnetic resonance (EPR), and time-resolved optical techniques. Several lines of EPR evidence suggested that the forward electron transfer rate to Q(A), k(Q), was slowed in those strains containing altered Q(A) sites. The slower rates of secondary electron transfer were confirmed by time-resolved optical results with the M246Glu-M247Asp mutations in the Q(A) site resulting in a dramatically lowered secondary electron transfer efficiency [k(Q) <(2 ns)-1] in comparison with either the native R. capsulatus RC or the Q(B) site mutant [k(Q) ≃ (200 ps)-1]. Secondary electron transfer in the two revertants was intermediate between that of the native RC and the Q(A) mutant. The P+Q(A)- → PQ(A) charge recombination rates were also changed in the strains that carried altered Q(A) sites. We show that local mutations in the Q(A) site, presumably through local electrostatic changes, significantly alter binding and electron transfer properties of Q(A).
KW - Electron paramagnetic resonance
KW - Electron spin polarization
KW - Electrostatics
KW - Quinone binding
KW - Revertant isolation
UR - http://www.scopus.com/inward/record.url?scp=8544219694&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=8544219694&partnerID=8YFLogxK
U2 - 10.1023/A:1005806700335
DO - 10.1023/A:1005806700335
M3 - Article
AN - SCOPUS:8544219694
VL - 52
SP - 93
EP - 103
JO - Photosynthesis Research
JF - Photosynthesis Research
SN - 0166-8595
IS - 2
ER -