Further Studies of Proton Translocations in Chloroplasts After Single-Turnover Flashes. II. Proton Deposition

Abstract

The deposition of protons in the inside spaces of pea class C chloroplasts was studied by means of the acidification of neutral red measured spectrophotometrically, with the outside space buffered.

Careful kinetic analysis of such signals revealed three components, during non-cyclic electron flow induced by single-turnover flashes. These components included a 'slow' phase not emphasized in previous studies. The half-times of these phases were: 'Fast', < 1 ms (not resolved); 'Intermediate', 13-25 ms with added electron acceptor or 4 ms without; and 'Slow', 70-90 ms.

Under conditions for cyclic electron flow only the I phase remained; it was the same magnitude as the I phase in non-cyclic flow, and its half-time was c. 3 ms. The F phase, which is usually attributed to protons from the oxidation of water, increased in average size with number of flashes (taken four flashes at a time) and was not fully patent until more than 20 flashes.

The size of the I phase, which is usually attributed to protons from the oxidation of plastohydroquinone, when measured in a sequence of flashes to dark-adapted suspensions under non- cyclic conditions, had a binary oscillation in phase with the oscillation in proton uptake reported previously. It was concluded that protons leave PQH₂ two at a time on alternate flashes.

The S phase (average in 10 test flashes) was reduced by fast preflashes; an origin near photosystem II is suggested. The S phase may imply a small pool of proton-sequestering ability near the water oxidation site, or a number of other possibilities.

In steady-state conditions, the ratio of the protons from PQH₂ to those from water was 1.0 under all conditions examined except in the absence of added electron acceptor, when it was as high as 1.6. This was the only condition apparently indicating a Q-cycle, with infrequent single-turnover flashes.