Assay of photoinhibition of photosystem II and protease activity.

Under light-stress conditions, the photosystem (PS)II reaction center D1 protein is photo-damaged. The damage to the D1 protein is induced by singlet oxygen molecules and endogenous free radicals generated by the photochemical reactions of PSII. To maintain PSII activity, the oxidatively damaged D1 protein is replaced by a newly synthesized protein. Thus, degradation and removal of the photodamaged D1 protein in PSII are essential steps for maintaining the viability of PSII. In the present chapter, we describe the method to induce photoinhibition of PSII both in vitro and in vivo, and also the method to assay the processes closely related to the photoinhibition, including degradation of the damaged D1 protein and its crosslinking with the neighboring polypeptides. The method to analyze the protease activity in the stroma that recognizes and digests the crosslinked products of the D1 protein generated by the light stress is also described.