PsaC | PSI-C core subunit of photosystem I (PsaC antibody + PsaC protein positive control )
AS04 042P | Clonality: Polyclonal | Host: Rabbit | Reactivity: [global antibody] for higher plants, algae, cyanobacteria
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KLH-conjugated synthetic peptide conserved in all known PsaC proteins includingArabidopsis thaliana AtCg01060, Hordeum vulgare P69416, Oryza sativa P0C360, Chlamydomonas reinhardtii Q00914, Synechococcus elongatus Q31QV2
Algae, Cyanobacteria, Glycine max, Nicotiana tabacum, Physcomitrium patens, Prochlorococcus sp. (surface and a deep water ecotype), Spinacia oleracea
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2 µg of total protein from Horderum vulgare (1), Chlamydomonas reinhardtii total cell (2), Synechococcus sp. 7942 total cell (3), were extracted with PEB (AS08 300) and separated on 4-12% NuPage (Invitrogen) LDS-PAGE and blotted 1h to PVDF. Blots were blocked immediately following transfer in 2% blocking reagent (GE Healthcare) in 20 mM Tris, 137 mM sodium chloride pH 7.6 with 0.1% (v/v) Tween-20 (TBS-T) for 1h at room temperature with agitation. Blots were incubated in the primary antibody at a dilution of 1: 50 000 for 1h at room temperature with agitation. The antibody solution was decanted and the blot was rinsed briefly twice, then washed once for 15 min and 3 times for 5 min in TBS-T at room temperature with agitation. Blots were incubated in secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated) diluted to 1:10 000 in 2% blocking solution for 1h at room temperature with agitation. The blots were washed as above and developed for 5 min with chemiluminescent detection reagent according the manufacturers instructions. Images of the blots were obtained using a CCD imager (FluorSMax, Bio-Rad) and Quantity One software (Bio-Rad).
Reactant: Arabidopsis thaliana (Thale cress)
Application: Western Blotting
Pudmed ID: 32269582
Journal: Front Plant Sci
Figure Number: 7B
Published Date: 2020-04-10
First Author: Pralon, T., Collombat, J., et al.
Impact Factor: 5.435Open Publication
Double mutant maintains thylakoid protein phosphorylation and state transitions after high light. (A) Total protein extracts of wild type (WT), abc1k1.1, -2, abc1k3.1, -2, and abc1k1/abc1k3.1, -2 light-exposed leaves were separated by SDS PAGE, transferred on nitrocellulose membrane and decorated with anti-phosphothreonine antibody. The main thylakoid phospho-proteins are indicated on the right according to their size. Core photosystem II proteins D1 (PsbA) and D2 (PsbD) are indicated together due their poor resolution. (B) The accumulation of the principal photosynthetic complexes was assessed using antibodies against specific subunits of each complex: anti-Lhcb2 for the major LHCII, anti-D1 (PsbA) for PSII, anti-PetC for cytochrome b6f, anti-PsaD and anti-PsaC for PSI, and anti-AtpC for ATP synthase. Actin signal is shown as a loading control. (C) Fluorescence quenching related to the state transitions (qT) of wild type (WT), abc1k1.1, -2, abc1k3.1, -2, and abc1k1/abc1k3.1, -2 under moderate light (120 ?mol of photons m–2 s–1) (ML) and after 3 h of high light (500 ?mol of photons m–2 s–1) (HL). qT was calculated from the maximal chlorophyll fluorescence measured after 10 min exposure to red light (660 nm) supplemented with far-red illumination (720 nm) “State 1” (FMST1) or to pure red light “State 2” (FMST2). Quenching related to state transition was calculated as qT = (FMST1 – FMST2)/FM. Each value represents the average of a pot containing 2–3 plants. Superscript letters are used to indicate statistically different groups (p < 0.05) by paired Student’s t-test.
Peptide target used to elicit this antibody is well conserved in all photoautotrophs except some cyanobacteria, some red algae and Cyanophora paradoxa, which contain a conserved substitution of a valine to an isoleucine. The performance of the antibodies has been confirmed against taxa containing both the valine and isoleucine variants.
For reconstitution of PsaC antibodies (AS10 939) add 50 µl of sterile water. For reconstitution of PsaC positive control add 100 µl of sterile water.
In some species minor cross reactions with some larger proteins are seen. These may contain related iron-sulfur binding motifs. Therefore size verification of the reacting band is required. Due to the small size of the protein, care should be taken to differentiate between chemiluminescent signal from PsaC and non-specific signals from chlotophylls or lipids if pigment is retained near the bottom of the blot.
Protein standard: use a load of 2 µl per well with ECL detection system and 4 µl per well with alkaline phospatase.
PsaC is a conserved, chloroplast-encoded, Fe-S binding protein of approximately 10 kDa, present in all known Photosystem I complexes. It is located on the stromal side of the thylacoid membranes. PsaC coordinates the Fe–S clusters FA and FB through two cysteine-rich domains.
Oesterhelt et al (2007). Regulation of photosynthesis in the unicellular acidophilic red alga Galdieria sulphuraria. Plant J.3:500511.
Ifuku et al. (2005). PsbP protein, but not PsbQ protein, is essential for the regulation and stabilization of photosystem II in higher plants. Plant Physiol. 3:1175-1184.
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