Anti-PSB33 | Rieske (2Fe-2S) domain-containing protein

Western blot analysis of selected candidate proteins (AtpB, 2Fe-2S, GLN2, Psb R, PRK, ALD) with their corresponding molecular weight in rice leaves during As stress under various S regimes (A); RuBisCo large subunit a band in coomassie blue staining (CBB) SDS gel served as a loading control (B), which used for normalization of detected proteins in densitometry studies (C) fold change indicated with respect of control samples.

PSB33 protein is mainly located in non-appressed thylakoids. Digitonin-solubilized thylakoid fractions (stroma lamellae (SL), grana margins (GM), and grana (G)) from wild-type plants acclimated to dark, low light (50 PFD), and high light (800 PFD) for 2 h were separated on an SDS gel, transferred to a nitrocellulose membrane and subsequently immunodecorated with an antibody against PSB33. A total protein stain (Ponceau) of the membrane shows that the same amount of protein was loaded and a clear protein-specific pattern of fractionation.

Reversible PSII-LHCII phosphorylation is defective in psb33 plants in fluctuating light conditions. Dynamics of thylakoid membrane phosphoproteins in response to fluctuating light conditions in wt and psb33. Thylakoids were isolated from plants at the end of the five dark–light periods (dark (Da1), low light for 2 h (LL1), high light for 2 h (HL), low light for 2 h (LL2), and darkness overnight (Da2)). On the left, a dilution series of dark (Da1) acclimated wild-type thylakoids shows the linear range of the antibody. (A) Immunoblot analyses of the PSII core (P-CP43, P-D2 and P-D1) and LHCII (P-LHCII) phosphoproteins from whole thylakoids were performed after SDS-PAGE. (B) Immunoblot showing the amounts of PsaB (above) and D1 (below) in the same samples as (A). Quantitative D1 protein levels, which have been normalized to Da1 D1 levels, are represented as numeric values below the D1 immunoblot. A Coomassie-stained PVDF membrane is shown as a loading control. (C) Immunoblots with specific antibodies against Lhcb1, P-Lhcb1, Lhcb2, P-Lhcb2, and PSB33 of whole-leaf protein extracts from wild-type and psb33 plants, highlighting the change in phosphorylation but not protein amounts of Lhcb1 and Lhcb2. A Coomassie-stained PVDF membrane is shown to demonstrate equal loading.