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pT360/pS364-β2 (phospho-β2-Adrenoceptor Antibody)

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  • 7TM0029B
  • 100 µl
  • Rabbit
Threonine360/Serine364 (T360/S364) is a major phosphorylation site of the β2 adrenoceptor.... more

Threonine360/Serine364 (T360/S364) is a major phosphorylation site of the β2 adrenoceptor. The pT360/pS364-β2 antibody detects phosphorylation in response to high- and low-efficacy agonists but not after PKC activation. T360/S364 phosphorylation is primarily mediated by GRK2 and is a key regulator of β2 desensitization, β-arrestin recruitment and internalization.

  Alternative Names β2,B2AR, ADRB2R, β2 Adrenergic... more

 

Alternative Names β2,B2AR, ADRB2R, β2 Adrenergic Receptor, β2-Adrenoceptor
IUPHAR Target ID  29
UniProt ID  P07550
Western Blot (WB)  1:1000
Immunocytochemistry (ICC) 1:100
Species Reactivity   Human
Host / Isotype Rabbit / IgG
Class Polyclonal
Immunogen A synthetic phosphopeptide derived from human β2 around the phosphorylation site of Thr360/Ser364.
Form  Liquid
Purification Antigen affinity chromatography
Storage buffer Dulbecco's PBS, pH 7.4, with 150 mM NaCl, 0.02% sodium azide
Storage conditions short-term 4°C, long-term -20°C
Figure 1. Agonist-induced Threonine360/Serine364 phosphorylation of the β2-Adrenoceptor.... more

Figure 1. Agonist-induced Threonine360/Serine364 phosphorylation of the β2-Adrenoceptor. Upper panel, HEK293 cells stably expressing the β2-Adrenoceptor (β2) were either not exposed or exposed to 1 µM of specific β2-adrenoceptor agonist Isoprenaline or 0.1 μM PMA (Phorbol 12-Myristate 13-Acetate) for 30 minutes. Cells were lysed and immunoblotted with the anti-pT360/pS364-β2 antibody (7TM0029B) at a dilution of 1:1000. Lower panel, blot was stripped and reprobed with the phosphorylation-independent anti-β2 antibody (7TM0029N) at a dilution of 1:1000 to confirm equal loading of the gel.

Figure 2. Analysis of time-dependent β2-Adrenoceptor phosphorylation using two phosphosite-specific antibodies. Upper two panels, HEK293 cells stably expressing the β2-Adrenoceptor (β2) were exposed to 1µM Isoprenaline for increasing periods of time ranging from 5 to 120 s. Cells were lysed and immunoblotted with the anti-pS355/pS356-β2 antibody (7TM0029A) or anti-pT360/pS364-β2 antibody (7TM0029B) at a dilution of 1:1000. Lower panel, blot was stripped and reprobed with the phosphorylation-independent anti-β2 antibody (7TM0029N) at a dilution of 1:1000 to confirm equal loading of the gel.

Figure 3. Immunocytochemical identification of Threonine360/Serine364 phosphorylation of β2-Adrenoceptor. Hela cells transiently expressing the FLAG-tagged  β2-Adrenoceptor (β2) were  exposed to 10 μM of specific β2-adrenoceptor agonist Isoprenaline for 5 min and immunocytochemically stained with an anti-FLAG antibody or the anti-pT360/pS364-β2 antibody (7TM0029B) at a dilution of 1:100.

Figure 4. Immunocytochemical identification of Threonine360/Serine364 phosphorylation of β2-Adrenoceptor. Hela cells transiently expressing the  β2-Adrenoceptor (β2) were either not exposed or exposed to 10 μM of specific β2-adrenoceptor agonist Isoprenaline and immunocytochemically stained with the anti-pT360/pS364-β2 antibody (7TM0029B) at a dilution of 1:100. Note, Threonine360/Serine364-phosphorylated β2 receptors were not detectable in untreated cells (0 min). Threonine360-phosphorylated β2 receptors were seen at the plasma membrane after 5 min Isoprenaline stimulation.

 

Benkel T, Zimmermann M, Zeiner J, Bravo S, Merten N, Lim VJY, Matthees ESF, Drube J,... more

Benkel T, Zimmermann M, Zeiner J, Bravo S, Merten N, Lim VJY, Matthees ESF, Drube J, Miess-Tanneberg E, Malan D, Szpakowska M, Monteleone S, Grimes J, Koszegi Z, Lanoiselée Y, O'Brien S, Pavlaki N, Dobberstein N, Inoue A, Nikolaev V, Calebiro D, Chevigné A, Sasse P, Schulz S, Hoffmann C, Kolb P, Waldhoer M, Simon K, Gomeza J, Kostenis E. How Carvedilol activates β2-adrenoceptors. Nat Commun. 2022 Nov 19;13(1):7109. doi: 10.1038/s41467-022-34765-w. PMID: 36402762; PMCID: PMC9675828.

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