Photoreduction of the secondary Photosystem I electron acceptor vitamin K1 in intact spinach chloroplasts and Cyanobacteria in vivo

AbstractThe light-induced EPR changes in the g=2 region of intact spinach chloroplasts and of marine cyanobacteria Synechococcus PCC 7002 at room temperature have been analysed. Illumination after dark adaptation results in the well known transient oxidation of P700 and its subsequent re-reduction during the first 100 ms of the illumination, as monitored by EPR Signal I. Here we show, that after a distinct lag phase, coinciding with the re-reduction of P700, a second EPR signal is generated. This signal could be identified as the reduction signal of phylloquinone electron acceptor A1 by its asymmetric spectrum (g=2.0049, ΔHptp=1.00 mT) and by its sensitivity to conditions which suppress limitation of electron transport at the acceptor side of PS I during illumination. Since steady state reduction of A1 has to date been found only by photoaccumulation in prereduced PS I particles at cryogenic temperatures, this is the first observation of the species under physiological conditions. A1 reduction is shown to saturate with light intensity at a level of 0.5 per P700. The observation of substantial levels of reduced A1 in vivo should provide new insights into the mechanisms of energy dissipation and light regulation of the reaction centre of PS I under acceptor limited conditions

Continuous ECS-indicated recording of the proton-motive charge flux in leaves

Technical features and examples of application of a special emitter–detector module for highly sensitive measurements of the electrochromic pigment absorbance shift (ECS) via dual-wavelength (550–520 nm) transmittance changes (P515) are described. This device, which has been introduced as an accessory of the standard, commercially available Dual-PAM-100 measuring system, not only allows steady-state assessment of the proton motive force (pmf) and its partitioning into ΔpH and ΔΨ components, but also continuous recording of the overall charge flux driven by photosynthetic light reactions. The new approach employs a double-modulation technique to derive a continuous signal from the light/dark modulation amplitude of the P515 signal. This new, continuously measured signal primarily reflects the rate of proton efflux via the ATP synthase, which under quasi-stationary conditions corresponds to the overall rate of proton influx driven by coupled electron transport. Simultaneous measurements of charge flux and $CO_2$ uptake as a function of light intensity indicated a close to linear relationship in the light-limited range. A linear relationship between these two signals was also found for different internal $CO_2$ concentrations, except for very low $CO_2$, where the rate of charge flux distinctly exceeded the rate of CO2 uptake. Parallel oscillations in $CO_2$ uptake and charge flux were induced by high $CO_2$ and $O_2$. The new device may contribute to the elucidation of complex regulatory mechanisms in intact leaves