SELECTIVE DESTRUCTION OF IRON-SULFUR CENTERS BY HEAT ETHYLENE-GLYCOL TREATMENT AND ISOLATION OF PHOTOSYSTEM-I CORE COMPLEX

Heat treatment of spinach thylakoids or Photosystem I particles in the presence of ethylene glycol caused the selective destruction of iron sulfur centers FA, FB and FX as judged by low-temperature EPR and flash spectroscopy. Incubation of thylakoids or Photosystem I particles in the presence of 50% ethylene glycol for 5 min selectively destroyed FA and FB at temperatures between 50 and 55°C, and FX between 60 and 70°C. P-700 was destroyed only above 70°C, and electron transport from ascorbate-dichlorophenolindophenol to methyl viologen was inhibited between 35 and 50°C by the same treatment. The destruction of FA and FB occurred at the same temperature range and resulted in the decrease of the decay phase with a 30 ms half-time in re-reduction kinetics of flash-oxidized P-700. This was accompanied by the appearance of the 1 ms decay phase, which reflects reduction of P-700+ by F- X. The 1 ms phase disappeared when FX was destroyed. Photosystem I core complex was isolated by solubilizing the heat/ethylene glycol-treated Photosystem I particles with Triton X-100, followed by sucrose density gradient ultracentrifugation. The complex obtained from 60°C/ethylene glycol-treated preparation lacked the 8 kDa FA/FB polypeptide and was composed of the polypeptides of apparent molecular weight 63, 60 and 5 kDa. This complex showed a small and broadened g = 1.77 FX signal. It is concluded that the heat/ethylene glycol treatment gives a simple and selective degradation-isolation method of Photosystem I reaction center complex. © 1990.