ACVR2A His Tag Protein, Human

ACVR2,ACTRII,activin A receptor type II

Datasheet COA

价格￥2,200.00 （供应商现货： 3-5个工作日）

货号 UA085066

规格

10μg50μg100μg500μg

数量

产品介绍

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产品规格

物种

Human
分子别名

ACVR2, ACTRII, activin A receptor type II
Accession

P27037
表达序列

Pro191-Ile488 with His Tag at the N-Terminus
表达宿主

Baculovirus-InsectCells
分子量

35-40kDa (Reducing)
纯度

＞95% by SDS-PAGE
活性

The specific activity of ACVR2A His Tag Protein was determined to be >7 pmol/min/ug as per activity assay protocol.
标记

Unconjugated
标签

His Tag
性状

Liquid
缓冲体系

50mM Tris, 150mM NaCl, pH7.5, 1mM DTT, 10%Glycerol
储存条件

Stable for 12 months upon stored at -80℃ from the date of receipt. And avoid repeated freeze-thaws cycles.
文献引用

1. Lee, S.J., et al. (2005). Regulation of muscle growth by multiple ligands signaling through activin type II receptors. Proceedings of the National Academy of Sciences, 102(50), 18117-18122.
2. Olaru, A., et al. (2003). Loss of heterozygosity and mutational analyses of the ACTRII gene locus in human colorectal tumors. Laboratory Investigation, 83(12), 1867-1871.
3. Rossi, M.R., et al. (2005). Truncating mutations in the ACVR2 gene attenuates activin signaling in prostate cancer cells. Cancer Genetics and Cytogenetics, 163(2), 123-129.

背景介绍

ACVR2A (Activin A Receptor Type 2A) is a transmembrane serine/threonine kinase receptor that serves as a type II receptor for the TGF-β superfamily. It is encoded by the gene located on chromosome 2q22.3 and is expressed in multiple tissues including brain, testis, intestine, liver, and kidney.
Mechanism of Action: ACVR2A functions as a constitutively active kinase that binds ligands including activin A, activin B, inhibin A, GDF8/myostatin, and GDF11. Upon ligand binding, ACVR2A recruits and phosphorylates type I receptors (ACVR1B/ALK4, ACVR1/ALK2, or ACVR1C/ALK7) within their GS domains. The activated type I receptor then phosphorylates SMAD2 and SMAD3, which complex with SMAD4 and translocate to the nucleus to regulate gene transcription. Inhibitory SMAD7 can block this signaling cascade.

操作步骤

实验方法
实验原理：使用ADP-Glo™激酶检测试剂盒进行ACVR2A活性测定，该试剂盒用于定量检测ACVR2A反应产生的ADP量。具体步骤如下：首先加入ADP-Glo™试剂以终止激酶反应并消耗剩余ATP；随后加入激酶检测试剂，将ADP转化为ATP，并通过荧光素酶/荧光素反应体系检测新合成的ATP。
实验材料
1.Kinase assay buffer(5X): 200 mM Tris-HCl, pH 7.4, 100 mM MgCl₂ and 0.5 mg/mL BSA, 250 μM DTT
2.Kinase assay buffer(1X): 40 mM Tris-HCl, pH 7.4, 20 mM MgCl₂, 0.1 mg/mL BSA, 50 μM DTT
3.ACVR2A His Tag Protein, Human
4.ADP-Glo Kinase Assay (UA, Catalog # UA070101)
5.底物: MBP Protein (Sinobiological, Catalog # M42-51N)
6.Solid white multi-well plate (384-well plate) (Corning, Catalog #3572)
7.读板器（PerkinElmer）
实验步骤
1.配制底物/ATP混合液，操作如下（以25 μM为例）：
Sample Name
Amount (μL)
10 mM ATP Solution
1
Kinase Assay Buffer III (5x)
79
Substrate at 0.5 mg/mL
80

2.用激酶检测缓冲液（1x）将ACVR2A稀释至40 µg/mL, 20 µg/mL和10 µg/mL，并在 384 孔板的每个孔中加入3 µL。
3.通过向每个孔中加入2 µL微升步骤1中配制的检测系统来启动反应。设置一个仅加入 3 µL激酶检测缓冲液（1x）的检测系统作为空白对照。反应总体积为5 µL。将反应在室温（22–25℃）下孵育40分钟。
4.向反应完成的体系中加入5 µL ADP-Glo Reagent，短暂混匀后在室温（22–25℃）下孵育 40分钟。
5.加入10 µL Detection Reagent，将板在室温（22–25℃）下孵育30分钟。
6.分别使用终点模式读取化学发光信号。
7.计算比活性。
标准曲线
1.在激酶检测缓冲液（1×）中将ATP和ADP稀释至25 μM。
2.按下表所示，混合25 μM ATP和25 μM ADP，配制ATP+ADP混合液，并在384孔板的每个孔中加入5 μL。
Well Number
1
2
3
4
5
6
7
8
9
10
11
12
25μM ADP (μL)
100
80
60
40
20
10
5
4
3
2
1
0
25μM ATP (μL)
0
20
40
60
80
90
95
96
97
98
99
100

3.向反应完成的孔中加入5 μL ADP-Glo试剂，短暂混匀，在室温（22-25°C）下孵育40分钟。
4.加入10 μL检测试剂，在室温（22-25°C）下孵育30分钟。
5.在终点模式下分别读取化学发光信号。
6.检测光信号并建立转换曲线。
Specific Activity (pmol/min/μg) =

ATP (pmol)-Blank

Incubation time(min) ×amount of enzyme (μg)

生物活性
- The specific activity of ACVR2A His Tag Tag Protein was determined to be >7 pmol/min/ug as per activity assay protocol.
电泳
- 2μg (R: reducing condition, N: non-reducing condition).