Recruitment of a foreign quinone into the A1 site of photosystem I. Altered kinetics of electron transfer in phylloquinone biosynthetic pathway mutants studied by time-resolved optical, EPR, and electrometric techniques.
Interruption of the menA or menB gene in Synechocystis sp. PCC 6803 results in the incorporation of a foreign quinone, termed Q, into the A(1) site of photosystem I with a number of experimental indicators identifying Q as plastoquinone-9. A global multiexponential analysis of time-resolved optical spectra in the blue region shows the following three kinetic components: 1) a 3-ms lifetime in the absence of methyl viologen that represents charge recombination between P700(+) and an FeS(-) cluster; 2) a 750-microseconds lifetime that represents electron donation from an FeS(-) cluster to methyl viologen; and 3) an approximately 15-microseconds lifetime that represents an electrochromic shift of a carotenoid pigment. Room temperature direct detection transient EPR studies of forward electron transfer show a spectrum of P700(+) Q(-) during the lifetime of the spin polarization and give no evidence of a significant population of P700(+) FeS(-) for t </= 2-3 microseconds. The UV difference spectrum measured 5 microseconds after a flash shows a maximum at 315 nm, a crossover at 280 nm, and a minimum at 255 nm as well as a shoulder at 290-295 nm, all of which are characteristic of the plastoquinone-9 anion radical. Kinetic measurements that monitor Q at 315 nm show a major phase of forward electron transfer to the FeS clusters with a lifetime of approximately 15 microseconds, which matches the electrochromic shift at 485 nm of the carotenoid, as well as an minor phase with a lifetime of approximately 250 microseconds. Electrometric measurements show similar biphasic kinetics. The slower kinetic phase can be detected using time-resolved EPR spectroscopy and has a spectrum characteristic of a semiquinone anion radical. We estimate the redox potential of plastoquinone-9 in the A(1) site to be more oxidizing than phylloquinone so that electron transfer from Q(-) to F(X) is thermodynamically unfavorable in the menA and menB mutants.
Semenov
A Y
AY
A. N. Belozersky Institute of Physicochemical Biology, Moscow State University, 119899 Moscow, Russia.
Vassiliev
I R
IR
van Der Est
A
A
Mamedov
M D
MD
Zybailov
B
B
Shen
G
G
Stehlik
D
D
Diner
B A
BA
Chitnis
P R
PR
Golbeck
J H
JH
D019883:Alkyl and Aryl Transferases / Q000235:genetics; D002734:Chlorophyll / Q000378:metabolism; D000458:Cyanobacteria / Q000235:genetics / Q000378:metabolism* / Q000528:radiation effects; D004578:Electron Spin Resonance Spectroscopy; D004579:Electron Transport; D005609:Free Radicals; D007700:Kinetics; D009154:Mutation; D010084:Oxidation-Reduction; D045322:Photosynthetic Reaction Center Complex Proteins / Q000235:genetics / Q000378:metabolism* / Q000528:radiation effects; D010971:Plastoquinone / Q000378:metabolism*; D011199:Potentiometry; D013053:Spectrophotometry / Q000379:methods; D013816:Thermodynamics; D010837:Vitamin K 1 / Q000378:metabolism*
D005609:Free Radicals; D045322:Photosynthetic Reaction Center Complex Proteins; D002734:Chlorophyll; C001785:chlorophyll P 700; D010837:Vitamin K 1; D019883:Alkyl and Aryl Transferases; C412723:phytyltransferase; D010971:Plastoquinone