Electron transfer in cyanobacterial photosystem I: I. Physiological and spectroscopic characterization of site-directed mutants in a putative electron transfer pathway from A0 through A1 to FX.
The Photosystem I (PS I) reaction center contains two branches of nearly symmetric cofactors bound to the PsaA and PsaB heterodimer. From the x-ray crystal structure it is known that Trp697PsaA and Trp677PsaB are pi-stacked with the head group of the phylloquinones and are H-bonded to Ser692PsaA and Ser672PsaB, whereas Arg694PsaA and Arg674PsaB are involved in a H-bonded network of side groups that connects the binding environments of the phylloquinones and FX. The mutants W697FPsaA, W677FPsaB, S692CPsaA, S672CPsaB, R694APsaA, and R674APsaB were constructed and characterized. All mutants grew photoautotrophically, yet all showed diminished growth rates compared with the wild-type, especially at higher light intensities. EPR and electron nuclear double resonance (ENDOR) studies at both room temperature and in frozen solution showed that the PsaB mutants were virtually identical to the wild-type, whereas significant effects were observed in the PsaA mutants. Spin polarized transient EPR spectra of the P700+A1- radical pair show that none of the mutations causes a significant change in the orientation of the measured phylloquinone. Pulsed ENDOR spectra reveal that the W697FPsaA mutation leads to about a 5% increase in the hyperfine coupling of the methyl group on the phylloquinone ring, whereas the S692CPsaA mutation causes a similar decrease in this coupling. The changes in the methyl hyperfine coupling are also reflected in the transient EPR spectra of P700+A1- and the CW EPR spectra of photoaccumulated A1-. We conclude that: (i) the transient EPR spectra at room temperature are predominantly from radical pairs in the PsaA branch of cofactors; (ii) at low temperature the electron cycle involving P700 and A1 similarly occurs along the PsaA branch of cofactors; and (iii) mutation of amino acids in close contact with the PsaA side quinone leads to changes in the spin density distribution of the reduced quinone observed by EPR.
Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, Iowa 50011, USA.
Wu
W
Xu
Parag
P
Chitnis
Alfia
A
Valieva
Art
A
van der Est
Yulia N
YN
Pushkar
Maciej
M
Krzystyniak
Christian
C
Teutloff
Stephan G
SG
Zech
Robert
R
Bittl
Dietmar
D
Stehlik
Boris
B
Zybailov
Gaozhong
G
Shen
John H
JH
Golbeck
D001665:Binding Sites; D015153:Blotting, Western; D002734:Chlorophyll / Q000737:chemistry; D018360:Crystallography, X-Ray; D000458:Cyanobacteria / Q000502:physiology*; D019281:Dimerization; D004578:Electron Spin Resonance Spectroscopy; D004579:Electron Transport; D004583:Electrons; D007700:Kinetics; D008027:Light; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010100:Oxygen / Q000378:metabolism; D045322:Photosynthetic Reaction Center Complex Proteins / Q000235:genetics* / Q000378:metabolism*; D020161:Physical Chromosome Mapping; D017354:Point Mutation; D011485:Protein Binding; D011809:Quinones / Q000737:chemistry; D013053:Spectrophotometry; D013696:Temperature; D020524:Thylakoids / Q000378:metabolism; D013997:Time Factors
D045322:Photosynthetic Reaction Center Complex Proteins; D011809:Quinones; D002734:Chlorophyll; D010100:Oxygen