# The Functions of Chloroplastic Ascorbate in Vascular Plants and Algae
## 5 Ascorbate May Impair the Oxygen-Evolving Complex  
When considering the physiological roles of Asc, it has to be taken into account that it is a reductant, and therefore, its cellular concentration is to be maintained in a particular range [114]. The basal Asc concentration in green algae is very low compared to higher plants (approx. 60 to 100 \(\upmu\)M in _C. reinhardtii_ and 5 mM in Arabidopsis) [74; 75; 77; 115]. It was observed that upon sulfur deprivation of _C. reinhardtii_, Asc accumulates to the mM range and that, in this range, Asc over-reduces the Mn cluster of OEC [76; 77]. The exact mode of action, i.e., as to which S-state is being inactivated, is not understood. Once the Mn-cluster is over-reduced by Asc, it may continuously provide electrons to Tyrz\({}^{+}\). However, the electron donation by Asc to PSII is relatively slow (halftime of approx. 20 to 50 ms, [99]) in comparison with the rate of electron transfer from intact OECs to Tyrz\({}^{+}\) (halftime of about 0.1 to 1 ms; [116]); for this reason, Asc cannot entirely prevent the accumulation of Tyrz\({}^{+}\) and P680\({}^{+}\) upon illumination in sulfur-deprived _C. reinhardtii_ cultures. Furthermore, strongly oxidizing species lead to donor-side induced photoinhibition, resulting in the relatively rapid degradation of PSII reaction center proteins, including PsbA, CP43, PSBO, and possibly others [77]. The loss of PSII activity could be regarded as damage induced by sulfur deprivation. However, upon downregulation of PSII activity, overexcitation and further photodamageare minimized. The metabolic changes downregulating photosynthetic activity and cell proliferation may serve to preserve cellular sulfur content and avoid more substantial damage [77]. On the other hand, the inactivation of OECs contributes to the establishment of hypoxia enabling hydrogenase expression, and H\({}_{2}\) production will act as a safety valve for photosynthetic electron transport [117]. Thereby, the damage imposed by sulfur limitation is minimized, and the alga cells may recover if sulfur becomes available again [118]. Intriguingly, Asc inactivates the OEC in green algae when accumulated to the mM range, but the same Asc concentration in vascular plants is physiological [115], which enables a full operation of OEC activity. On the other hand, it was demonstrated earlier that upon the chemical removal of the extrinsic OEC subunits in isolated PSII membranes, bulky reductants, including Asc, could directly reduce the Mn-cluster [119]. There are notable differences between the extrinsic OEC proteins of vascular plants and green algae [120; 121]. Namely, in green algae, the Mn-cluster is shielded by a 33 kDa PSBO subunit and two smaller subunits, PSBP and PSBQ, with some structural differences and binding properties in comparison to vascular plants [121; 122]. Moreover, in vascular plants, an additional subunit, PSBR, is found in the vicinity of the Mn-cluster [123]. The structural differences and the variance in binding properties of the extrinsic proteins may be key in explaining why Asc reduces the Mn-cluster in algae but not in higher plants when present in the same concentration range. It is also conceivable that, during evolution, as cellular Asc concentration increased [124], the extrinsic proteins have evolved to protect the OEC against the reducing effect of Asc present in the thylakoid lumen. In Arabidopsis subjected to darkness for 24 h, the Mn-cluster becomes inactivated, being one of the earliest effects of dark-induced senescence on photosynthesis [125]. Remarkably, the extent of OEC inactivation was much weaker in Asc-deficient mutants compared to wild-type plants, suggesting that Asc was responsible for the diminishment of oxygen evolution. In a _psbo1_ knockout mutant, the compromised OEC activity was further aggravated upon dark treatment, suggesting that the extrinsic proteins protect the OEC against the reducing effect of Asc.