Studies on the mechanism of ADRY agents (agents accelerating the deactivation reactions of water-splitting enzyme system Y) on thermoluminescence emission

Abstract

The effect of 2-(3-chloro-4-trifluoromethyl)anilino-3,5-dinitrothiophene (ANT-2p), known to be the most powerful ADRY agent (Renger, G. (1972) Biochim. Biophys. Acta 256, 428–439), on thermoluminescence has been investigated. Two thermoluminescence bands were analyzed: (a) the emission peaking at about 20–30°C caused by warming up of untreated chloroplasts, illuminated with a single 5 μs flash at room temperature and frozen rapidly to 77 K; and (b) the band emitted in the range of −10 up 10°C after warming of chloroplast suspensions containing 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) which were illuminated with a single 5 μs flash at −15°C and frozen rapidly at 77 K. These bands were attributed to the recombination of the B −S 2(S 3) and X-320 −S 2 states, respectively (Rutherford, A.W., Crofts, A.R. and Inoue, Y. (1982) Biochim. Biophys. Acta 682, 457–465). It was found that: (1) The B −S 2(S 3) band is markedly diminished at very low ANT-2p concentrations of less than one molecule per 2000 chlorophylls. (2) The inhibition of the X-320 −S 2 band requires significantly higher concentrations of ANT-2p (50% peak reduction at one ANT-2p molecule per 100 chlorophylls). (3) Preflashing at room temperature before cooling to −15°C diminishes the X-320 −S 2 band significantly in the presence of ANT-2p, while almost no effect is observed in its absence. (4) The state X-320 −S 2 decays monoexponentially with a half-lifetime of 2 min at −15°C in the absence of ANT-2p. In the presence of one ANT-2p molecule per 800 chlorophylls the decay becomes biphasic with half-lifetimes of 0.5 and 2 min and an amplitude ratio of 2:3, respectively. The results obtained can be explained consistently by the function of ANT-2p as an ADRY agent acting as a mobile species within the thylakoid membrane at room temperature. At subzero temperatures, a ‘fixed-place’ mechanism appears to be operative. The implications for the ADRY effect and thermoluminescence are discussed.