We used two different techniques to measure the recovery time of Photosystem II following the transfer of a single electron from P-680 to QA in thylakoid membranes isolated from spinach. Electron transfer in Photosystem II reaction centers was probed first by spectroscopic measurements of the electrochromic shift at 518 nm due to charge separation within the reaction centers. Using two short actinic flashes separated by a variable time interval we determined the time required after the first flash for the electrochromic shift at 518 nm to recover to the full extent on the second flash. In the second technique the redox state of QA at variable times after a saturating flash was monitored by measurement of the fluorescence induction in the absence of an inhibitor and in the presence of ferricyanide. The objective was to determine the time required after the actinic flash for the fluorescence induction to recover to the value observed after a 60 s dark period. Measurements were done under conditions in which (1) the electron donor for Photosystem II was water and the acceptor was the endogenous plastoquinone pool, and (2) Q400, the Fe2+ near QA, remained reduced and therefore was not a participant in the flash-induced electron-transfer reactions. The electrochromic shift at 518 nm and the fluorescence induction revealed a prominent biphasic recovery time for Photosystem II reaction centers. The majority of the Photosystem II reaction centers recovered in less than 50 ms. However, approx. one-third of the Photosystem II reaction centers required a half-time of 2-3 s to recover. Our interpretation of these data is that Photosystem II reaction centers consist of at least two distinct populations. One population, typically 68% of the total amount of Photosystem II as determined by the electrochromic shift, has a steady-state turnover rate for the electron-transfer reaction from water to the plastoquinone pool of approx. 250 e- / s, sufficiently rapid to account for measured rates of steady-state electron transport. The other population, typically 32%, has a turnover rate of approx. 0.2 e- / s. Since this turnover rate is over 1000-times slower than normally active Photosystem II complexes, we conclude that the slowly turning over Photosystem II complexes are inconsequential in contributing to energy transduction. The slowly turning over Photosystem II complexes are able to transfer an electron from P-680 to QA rapidly, but the reoxidation of Q-A is slow (t 1 2 = 2 s). The fluorescence induction measurements lead us to conclude that there is significant overlap between the slowly turning over fraction of Photosystem II complexes and PS IIβ reaction centers. One corollary of this conclusion is that electron transfer from P-680 to QA in PS IIβ reaction centers results in charge separation across the membrane and gives rise to an electrochromic shift.
ASJC Scopus subject areas
- Cell Biology