Functional and molecular characterization of plastid terminal oxidase from rice (Oryza sativa).

Abstract The plastid terminal oxidase (PTOX) is a plastohydroquinone:oxygen oxidoreductase that shares structural similarities with alternative oxidases (AOX). Multiple roles have been attributed to PTOX, such as involvement in carotene desaturation, a safety valve function, participation in the processes of chlororespiration and setting the redox poise for cyclic electron transport. We have investigated a homogenously pure MBP fusion of PTOX. The protein forms a homo-tetrameric complex containing 2 Fe per monomer and is very specific for the plastoquinone head-group. The reaction kinetics were investigated in a soluble monophasic system using chemically reduced decyl-plastoquinone (DPQ) as the model substrate and, in addition, in a biphasic (liposomal) system in which DPQ was reduced with DT-diaphorase. While PTOX did not detectably produce reactive oxygen species in the monophasic system, their formation was observed by room temperature EPR in the biphasic system in a [DPQH 2 ] and pH-dependent manner. This is probably the result of the higher concentration of DPQ achieved within the partial volume of the lipid bilayer and a higher Km observed with PTOX-membrane associates which is ≈ 47 mM compared to the monophasic system where a Km of ≈ 74 μM was determined. With liposomes and at the basic stromal pH of photosynthetically active chloroplasts, PTOX was antioxidant at low [DPQH 2 ] gaining prooxidant properties with increasing quinol concentrations. It is concluded that in vivo, PTOX can act as a safety valve when the steady state [PQH 2 ] is low while a certain amount of ROS is formed at high light intensities.