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 wa ve 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, t he continuous wave EPR spectrum of the photoaccumulated Q(-) radical, measu red 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 pro minent features: (i) Q(-) has a larger g-anisotropy than native phylloquino ne, (ii) Q(-) does not display the prominent methyl hyperfine couplings att ributed to the a-methyl group of phylloquinone, and (iii) the orientation o f Q(-) in the A(1) site as derived from the spin polarization is that of na tive phylloquinone in the wild type. Electron spin echo modulation experime nts on P700(+) Q(-) show that the dipolar coupling in the radical pair is t he same as in native PS I, i.e. the distance between P700(+) and Q(-) (25.3 +/- 0.3 Angstrom) is the same as between P700(+) and A(1)(-) in the wild t ype. Pulsed electron-nuclear double resonance studies show two sets of reso lved spectral features with nearly axially symmetric hyperfine couplings. T hey are tentatively assigned to the two methyl groups of the recruited plas toquinone-9, and their difference indicates a strong inequivalence among th e 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 orientatio n similar to that of phylloquinone in the wild type, and (iii) has spectros copic properties consistent with its identity as plastoquinone-9.