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 [DPQH₂] 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 ≈47mM compared to the monophasic system where a Km of ≈74μM was determined. With liposomes and at the basic stromal pH of photosynthetically active chloroplasts, PTOX was antioxidant at low [DPQH₂] gaining prooxidant properties with increasing quinol concentrations. It is concluded that in vivo, PTOX can act as a safety valve when the steady state [PQH₂] is low while a certain amount of ROS is formed at high light intensities.

The impact of photosynthesis on initiation of leaf senescence

“…Indeed, complex I besides complex III is regarded as the main source of these ROS in mitochondria (Chen et al , Murphy ). PTOX can also generate O 2 •‐ in a side reaction depending on substrate availability (Yu et al ), although this enzyme catalyzes as main reaction the direct reduction of O 2 to H 2 O. Two molecules of O 2 •‐ are converted to H 2 O 2 and O 2 by disproportionation, either spontaneously or catalyzed by superoxide dismutase (SOD).…”

Redox- and Reactive Oxygen Species-Dependent Signaling into and out of the Photosynthesizing Chloroplast

“…PTOX catalyzes the oxidation of plastoquinol and transfers the electrons to oxygen to form water. However, depending on the plastoquinol concentration, PTOX can generate O 2 $ 2 in a side reaction (Yu et al, 2014).…”

Distinguishing the roles of thylakoid respiratory terminal oxidases in the cyanobacterium Synechocystis sp. PCC 6803

“…However, since the PSI-less mutant is highly sensitive to light (Shen et al, 1993), the contribution of Cox as an electron shuttle to oxygen is likely to be less efficient compared with PSI activity, or this could be a transient phenomenon. It is possible that, under specific conditions, Cox also produces reactive oxygen species via a mechanism similar to PTOX in plants (Heyno et al, 2009;Feilke et al, 2014;Yu et al, 2014), thereby generating oxidative damage to the cells. On the other hand, rapid light curve analysis demonstrated a slight but significant difference in Y(ND) values between Δcyd and Δcox/cyd cells under higher light intensities, indicating competition between PSI and Cox for electrons in the Δcyd mutant (Fig.…”