AtpB | Beta subunit of ATP synthase (chloroplastic + mitochondrial) (chicken)
AS03 030 | Clonality: Polyclonal | Host: Chicken | Reactivity: [global antibody] for plant and bacterial F-type ATP synthases
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KLH-conjugated synthetic peptide derived from available plant, algal (chloroplastic and mitochondrial) and bacterial sequences of beta subunits of F-type ATP synthases, including Arabidopsis thaliana chloroplastic ATP synthase subunit beta UniProt: P19366, TAIR: AtCg00480 and Arabidopsis thaliana mitochondrial ATP synthase subunit beta-1, UniProt: P83483, TAIR: At5g08670 as well as Chlamydomonas reinhardtii, UniProt: P06541 and A8IQU3
53.9 kDa (Arabidopsis thaliana), 51.7 kDa (Synechocystis PCC 6803), 53.7 kDa (Spinacia oleracea)
10 µg of total protein from samples such as beef muscle (1), rat liver (2), Arabidopsis thaliana leaf (3), Hordeum vulgare leaf (4), Synechocystis PCC 6803 total cell (5), Synechococcus PCC 7942 total cell (6) were extracted with Protein Extraction Buffer PEB (AS08 300). Samples were diluted with 1X sample buffer (NuPAGE LDS sample buffer (Invitrogen) supplemented with 50 mM DTT and heat at 70C for 5 min and keep on ice before loading. Protein samples were separated on Bolt 4-12% Tris-Bis gel (Invitrogen) LDS-PAGE and blotted for 1h to 1.5h on PVDF using Bolt Mini Blot Module. Blot was blocked immediately following transfer in 2% blocking reagent (GE RPN 2125; Healthcare) dissolved 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. Blot was incubated in the primary antibody at a dilution of 1: 5 000 (in blocking reagent) for 1h at room temperature with agitation. The antibody solution was decanted and the blot was rinsed briefly twice, and then washed 1x15 min and 3x5 min with TBS-T at room temperature with agitation. Blot was incubated in secondary antibody (anti-chicken IgY horse radish peroxidase conjugated) diluted to 1:20 000 in blocking reagent for 1h at room temperature with agitation. The blot was washed as above. Signals in the blot were detected using Lumigen ECL Ultra Reagent (Lumigen TMA-6, Lumigen), and visualized using the Molecular Imager VersaDoc MP 4000 System (Bio-Rad) and Quantity One software (Bio-Rad). Exposure time was 30 seconds.
The anti-AtpB antibody will detect the mitochondrial form of the F1 ATP synthase subcomplex, as well as the chloroplastic CF1 ATP synthase and most known bacterial F-type ATP synthases. Peptide used for antibody production is located in a beta sheet, which is partly exposed near the surface of the AtpB protein.
Results of immunogold studies using anti-AtpB antibody are published in Andersson et al. (2009).
ATP synthase is the universal enzyme that synthesizes ATP from ADP and phosphate using the energy stored in a transmembrane ion gradient.
Quesada et al. (2011). Arabidopsis RUGOSA2 encodes an mTERF family member required for mitochondrion, chloroplast and leaf development. Plant J. Nov;68(4):738-53. doi: 10.1111/j.1365-313X.2011.04726.x. Epub 2011 Sep 13.
Andersson et. al (2009). Co-localization of P-glycerate kinase, P-ribulokinase, ADP-glucose pyrophosphorylase and Rubisco activase with CF1 in pea leaf chloroplasts. Plant Science 177:136-14. (immunologold)
Morash et al. (2007). Macromolecular dynamics of the photosynthetic system over a seasonal developmental progression in Spartina alterniflora. Canadian J. of Botany, 2007, 85(5): 476-483, 10.1139/B07-043.
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