Study on the photo-generation of superoxide radicals in Photosystem II with EPR spin trapping techniques.

Direct EPR evidence of the photo-generation of superoxide radicals (O(2) (-.)) was obtained by using a novel spin trapping probe in spinach Photosystem II (PS II) membrane fragments. The production of O(2) (-.) was detected by following the formation of 5-diethoxyphosphoryl-5-methyl-1-pyrroline-N-oxide (DEPMPO) superoxide adducts (DEPMPO-OOH). The inhibition of O(2) (-.) formation by 3-(3,4-dichlorophenyl) -1,1-dimethylurea (DCMU) and the 77 K fluorescence spectrum indicated that O(2) (-.) were generated from PS II, not from PS I. The inhibition of O(2) (-.) formation by DCMU also suggested that O(2) (-.) were generated from the Q(B)binding site, not at a site prior to DCMU blockage. The extrinsic proteins and Mn are very important to eliminate O(2) (-.), showing that the oxygen-evolving system is involved in O(2) (-.) removal rather than production.