ACVR2B Protein, Mouse (HEK293, His)

ACVR2B is a type II member of the TGF-β family of receptor Serine/Threonine kinases. ACVR2B binds to activins and growth differentiation factors (GDF), which in turn activate type I receptors, activating downstream molecule SMAD2/3^[1]. ACVR2B Protein, Mouse (HEK293, His) is produced in HEK293 cells with a C-Terminal His-tag. It consists of 134 amino acids (M1-T134).

Activin receptor type-2B (ACVR2B), also known as ActR-IIB and MGC116908, is an activin type II receptor. Type I receptors are essential for signaling; and type II receptors are required for binding ligands and for expression of type I receptors^[1].
The sequence of amino acids in ACVR2B proteins from different species is very stable, which leads to the conclusion that in the process of evolution, ACVR2B has been only slightly altered, and that both in humans and in animals, its function is similar.
Activins and growth differentiation factors (GDF) bind to ACVR2B, which in turn activate type I receptors such as activin receptor-like kinases (ALK) ALK4 and ALK5, activating downstream molecule SMAD2/3. SMADSs regulate a number of myogenic genes, such as myoD, myogenin, and Myf5, that are involved in cellular hypertrophy, proliferation, or differentiation. Noncanonical ACVR2B pathways have also been shown to regulate MAP kinases. ACVR2B blocks signaling of myostatin, its close homolog GDF11, as well as activin A, activin B, and BMP10^[1]. Activin A primarily binds to the type 1 receptors ALK4 or ALK7 in complex with ACVR2A or ACVR2B, causing activation of SMAD2 or SMAD3^[2].
In myeloma cells, BMP-6- and BMP-9-induced activation of SMAD1/5/8 through ACVR2A/ACVR2B/ALK2 is inhibited by activin A treatment^[2]. ACVR2B has been shown to preserve muscle mass and prolong survival in tumor hosts, and to increase bone mass in models of osteogenesis imperfecta and muscular dystrophy^[3].

Recombinant mouse ACVR2B (2 ng/mL; added 3 min before hypoxia) antagonizes SMAD2 signaling and cell death in cardiomyocytes that are subjected to hypoxic stress^[1].

Recombinant mouse ACVR2B (1 mg/kg; s.c.; once) shows cardioprotection activity in ischemia-reperfusion (IR) injury mouse. ACVR2B= modifies cardiac metabolism, left ventricular mitochondrial respiration, as well as cardiac phenotype toward physiological hypertrophy. By blocking myostatin, ACVR2B potentially reduces cardiomyocyte death and modifies cardiomyocyte metabolism for hypoxic conditions to protect the heart from IR injury^[1].

1.This product does not contain protein kinase domain.
2.Measured by its ability to neutralize Activin-mediated inhibition on MPC11 cell proliferation. The ED₅₀ for this effect is typically 0.3502 µg/mL in the presence of 10 ng/mL recombinant Activin A, corresponding to a specific activity is 2.856×10³ U/mg.

• 
  Measured by its ability to neutralize Activin-mediated inhibition on MPC11 cell proliferation. The ED₅₀ for this effect is typically 0.3502 µg/mL in the presence of 10 ng/mL recombinant Activin A, corresponding to a specific activity is 2.856×10³ U/mg.

Materials
ACVR2B Protein, Mouse (HEK293, His) (HY-P75536)
Cell line: MPC-11
Culture medium: DMEM + 10% HS + 1% P/S
Penicillinillin-Streptomycin (100×), Sterile (HY-K1006)
Cell Counting Kit-8 (CCK8 solution, HY-K0301)
Activin A Protein, Human/Mouse/Rat (HEK293, His) (HY-P73241)

Procedure
1. Seed cells in a 96-well plate at a density of 15,000 cells/well with a volume of 50 μL per well.
2. Prepare the protein in complete DMEM medium containing 10 ng/mL Activin A. Add 50 μL of complete medium containing different concentrations of Mouse ACVR2B protein to the 96-well plate, so that the final concentrations of ACVR2B protein in each well are 0, 0.0001, 0.001, 0.01, 0.05, 0.1, 0.2, 0.5, 1, 2, 5, 10, 50 μg/mL.
3. Incubate the cells in a 37℃, 5% CO₂ incubator for 72 h.
4. After the incubation time is reached, prepare complete medium containing CCK8 solution at a final concentration of 10% under light-protected conditions.
5. Add 10 μL of CCK8 solution to each well and let it react statically in the incubator for 4 h.
6. Measure the OD value at 450 nm using a microplate reader.
7. Use GraphPad Prism software to plot a curve with OD value as the ordinate and protein concentration as the abscissa, and calculate the ED₅₀ value.

Mouse

HEK293

C-His

P27040-1 (S19-T134)

11481  [NCBI]

Extracellular Domain

SGRGEAETRECIYYNANWELERTNQSGLERCEGEQDKRLHCYASWRNSSGTIELVKKGCWLDDFNCYDRQECVATEENPQVYFCCCEGNFCNERFTHLPEPGGPEVTYEPPPTAPT

23-37 kDa

• 
  ≥ 90%, as determined by reducing SDS-PAGE.

Lyophilized powder.

Lyophilized from a 0.22 μm filtered solution of PBS, pH 7.4.

<1 EU/μg, determined by LAL method.

It is not recommended to reconstitute to a concentration less than 100 μg/mL in ddH₂O. For long term storage it is recommended to add a carrier protein (0.1% BSA, 5% HSA, 10% FBS or 5% Trehalose).

Stored at -20°C for 2 years from date of receipt. After reconstitution, it is stable at 4°C for 1 week or -20°C for longer (with carrier protein). It is recommended to freeze aliquots at -20°C or -80°C for extended storage.

Room temperature in continental US; may vary elsewhere.

• [1]. Johanna Magga, et al. Systemic Blockade of ACVR2B Ligands Protects Myocardium from Acute Ischemia-Reperfusion Injury. Mol Ther. 2019 Mar 6;27(3):600-610.  [Content Brief]

  [2]. Oddrun Elise Olsen, et al. Activin A inhibits BMP-signaling by binding ACVR2A and ACVR2B. Cell Commun Signal. 2015 Jun 6;13:27.  [Content Brief]

  [3]. Rafael Barreto, et al. ACVR2B/Fc counteracts chemotherapy-induced loss of muscle and bone mass. Sci Rep. 2017 Oct 31;7(1):14470.  [Content Brief]