Lumen Thiol Oxidoreductase1, a Disulfide Bond-Forming Catalyst, Is Required for the Assembly of Photosystem II inArabidopsis   

Abstract

Here, we identify Arabidopsis thaliana Lumen Thiol Oxidoreductase1 (LTO1) as a disulfide bond-forming enzyme in the thylakoid lumen. Using topological reporters in bacteria, we deduced a lumenal location for the redox active domains of the protein. LTO1 can partially substitute for the proteins catalyzing disulfide bond formation in the bacterial periplasm, which is topologically equivalent to the plastid lumen. An insertional mutation within the LTO1 promoter is associated with a severe photoautotrophic growth defect. Measurements of the photosynthetic activity indicate that the lto1 mutant displays a limitation in the electron flow from photosystem II (PSII). In accordance with these measurements, we noted a severe depletion of the structural subunits of PSII but no change in the accumulation of the cytochrome b 6 f complex or photosystem I. In a yeast two-hybrid assay, the thioredoxin-like domain of LTO1 interacts with PsbO, a lumenal PSII subunit known to be disulfide bonded, and a recombinant form of the molecule can introduce a disulfide bond in PsbO in vitro. The documentation of a sulfhydryl-oxidizing activity in the thylakoid lumen further underscores the importance of catalyzed thiol-disulfide chemistry for the biogenesis of the thylakoid compartment.

PHOTOSYSTEM II PROTEIN33, a Protein Conserved in the Plastid Lineage, Is Associated with the Chloroplast Thylakoid Membrane and Provides Stability to Photosystem II Supercomplexes in Arabidopsis

“…3.63 6 0.97 -3.37 6 0.65 ns 5.64 6 1.00 -5.14 6 0.63 ns 7.04 6 1.18 -6.51 6 0.69 ns 7.64 6 0.86 -7.01 6 0.34 ns DC (170, 670, 1,270, 2,000 PPFD) 1.37 6 0.49 -2.39 6 0.80 ns 2.23 6 0.48 -2.10 6 0.50 ns 0.71 6 0.45 -0.63 6 0.63 ns ---PI 1.43 6 0.09 0.85 6 0.09* 0.62 6 0.09* Oxygen evolution 0.28 6 0.05 0.14 6 0.01* 0.13 6 0.01* which protons accumulate during active electron transport, and between the bacterial cytoplasm negative (n) side and the plastid stroma, which becomes proton deficient. High PhoA activity indicates a p-side location of the reporter, while LacZ activity indicates an n-side location of the reporter (Hamel et al, 2003;Karamoko et al, 2011). As shown in Supplemental Figure S3D, when the PhoA-LacZa reporter is fused to the N-terminal side of the transmembrane region, transformants display predominantly b-galactosidase activity, suggesting that the Rieske-like domain faces the stroma.…”

Activation of the Stt7/STN7 Kinase through Dynamic Interactions with the Cytochrome b6f Complex

“…This configuration of the kinase allows its catalytic domain to act on the substrate sites of the LHCII proteins, which are exposed to the stroma. Although in this model the conserved Cys residues in the lumen are on the opposite side from the stromal thioredoxins, it is possible that thiol-reducing equivalents are transferred across the thylakoid membrane through the CcdA and Hcf164 proteins, which have been shown to operate in this way during heme and Cyt b 6 f assembly (Lennartz et al, 2001;Page et al, 2004) or through the LTO1 protein (Du et al, 2015;Karamoko et al, 2011).…”

Thioredoxin and NADPH-Dependent Thioredoxin Reductase C Regulation of Tetrapyrrole Biosynthesis

“…In contrast, oxidized proteins with disulfide bridges or bound sulfenic acids cannot bind AMS. AMS-labeled proteins migrate more slowly on nonreducing SDS-PAGE gels than unlabeled proteins (Kobayashi et al, 1997;Ikegami et al, 2007;Karamoko et al, 2011;Wang et al, 2013). CHLM has three Cys residues (Cys-111, Cys-115, and Cys-177).…”