Recruitment of a foreign quinone into the A(1) site of photosystem I. II. Structural and functional characterization of phylloquinone biosynthetic pathway mutants by electron paramagnetic resonance and electron-nuclear double resonance spectroscopy.
Electron paramagnetic resonance (EPR) and electron-nuclear double resonance studies of the photosystem (PS) I quinone acceptor, A(1), in phylloquinone biosynthetic pathway mutants are described. Room temperature continuous wave EPR measurements at X-band of whole cells of menA and menB interruption mutants show a transient reduction and oxidation of an organic radical with a g-value and anisotropy characteristic of a quinone. In PS I complexes, the continuous wave EPR spectrum of the photoaccumulated Q(-) radical, measured at Q-band, and the electron spin-polarized transient EPR spectra of the radical pair P700(+) Q(-), measured at X-, Q-, and W-bands, show three prominent features: (i) Q(-) has a larger g-anisotropy than native phylloquinone, (ii) Q(-) does not display the prominent methyl hyperfine couplings attributed to the 2-methyl group of phylloquinone, and (iii) the orientation of Q(-) in the A(1) site as derived from the spin polarization is that of native phylloquinone in the wild type. Electron spin echo modulation experiments on P700(+) Q(-) show that the dipolar coupling in the radical pair is the same as in native PS I, i.e. the distance between P700(+) and Q(-) (25.3 +/- 0.3 A) is the same as between P700(+) and A(1)(-) in the wild type. Pulsed electron-nuclear double resonance studies show two sets of resolved spectral features with nearly axially symmetric hyperfine couplings. They are tentatively assigned to the two methyl groups of the recruited plastoquinone-9, and their difference indicates a strong inequivalence among the two groups when in the A(1) site. These results show that Q (i) functions in accepting an electron from A(0)(-) and in passing the electron forward to the iron-sulfur clusters, (ii) occupies the A(1) site with an orientation similar to that of phylloquinone in the wild type, and (iii) has spectroscopic properties consistent with its identity as plastoquinone-9.
Zybailov
B
B
Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park, Pennsylvania 16802, USA.
van der Est
A
A
Zech
S G
SG
Teutloff
C
C
Johnson
T W
TW
Shen
G
G
Bittl
R
R
Stehlik
D
D
Chitnis
P R
PR
Golbeck
J H
JH
D019883:Alkyl and Aryl Transferases / Q000235:genetics; D002734:Chlorophyll; D000458:Cyanobacteria / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D029968:Escherichia coli Proteins; D005609:Free Radicals; D006836:Hydro-Lyases / Q000235:genetics; D045342:Light-Harvesting Protein Complexes; D009154:Mutation; D045322:Photosynthetic Reaction Center Complex Proteins / Q000737:chemistry* / Q000378:metabolism; D045331:Photosystem I Protein Complex; D010971:Plastoquinone / Q000302:isolation & purification*; D010837:Vitamin K 1 / Q000096:biosynthesis
D029968:Escherichia coli Proteins; D005609:Free Radicals; D045342:Light-Harvesting Protein Complexes; D045322:Photosynthetic Reaction Center Complex Proteins; D045331:Photosystem I Protein Complex; D002734:Chlorophyll; C001785:chlorophyll P 700; D010837:Vitamin K 1; D019883:Alkyl and Aryl Transferases; C112258:MenA protein, E coli; D006836:Hydro-Lyases; C021853:naphthoate synthetase; D010971:Plastoquinone