Quantitative electron-paramagnetic-resonance measurements of the electron-transfer components of the photosystem-I reaction centre

Abstract

E.p.r. spectrometry was used to investigate the quantitative relationships between the oxidized chlorophyll free-radical signal I and the reduced iron-sulphur centre-A signal generated on illuminating Photosystem-I particles at cryogenic temperatures. In Photosystem-I particles prepared by using the French press or Triton X-100, at pH8.0 in the presence and absence of ascorbate and at pH 10.0 in the presence of ascorbate, the size of the light-induced signal I and iron-sulphur centre-A signals, corresponded to equal numbers of unpaired electron spins in each component. At 77K the spin-lattice relaxation time, T1, of the free radical signal I in samples of Photosystem-I particles prepared with Triton X-100 in the absence of ascorbate was 0.68 times the T1 value in the presence of ascorbate. Such changes in relaxation time can account for the different quantitative conclusions incorrectly arrived at from measurements made at saturating microwave powers [Bearden & Malkin (1976) Biochem. Biophys. Acta 430, 538-547; Malkin & Bearden (1976) FEBS Lett. 69, 216-220]. In the presence of benzoquinone and ferricyanide the ratio of free radical to centre A was 2.96:1, and at 77K the T1 was 0.50 times the T1 for ascorbate-treated samples. Here free radicals from bulk chlorophyll are generated in addition to those from the reaction-centre chlorophyll.