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产品中心 药物靶点和Fc受体 小分子药物靶点蛋白

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IDH1 His Tag Protein, Human

IDH1,PICD,IDP

价格 ￥600.00 （ 供应商现货 ： 3-5个工作日）
货号 UA085026
规格
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产品介绍

产品介绍 评论(0)

产品规格

  • 物种

    Human

  • 分子别名

    IDH1, PICD, IDP

  • Accession

    O75874-1

  • 表达序列

    Met 1 - Leu 414 with His Tag at the C-Terminus

  • 表达宿主

    E.coli

  • 分子量

    40-55kDa (Reducing)

  • 纯度

    ＞95% by SDS-PAGE,> 90% by HPLC

  • 活性

    Measured by the ability to oxidatively decarboxylate isocitrate to 2-oxoglutarate. The specific activity is >10000 pmol/min/μg, as measured under the described conditions.

  • 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. Parsons, D.W., et al. (2008). An integrated genomic analysis of human glioblastoma multiforme. Science, 321(5897), 1807-1812.
    2. Dang, L., et al. (2009). Cancer-associated IDH1 mutations produce 2-hydroxyglutarate. Nature, 462(7274), 739-744.
    3. Xu, W., et al. (2011). Oncometabolite 2-hydroxyglutarate is a competitive inhibitor of α-ketoglutarate-dependent dioxygenases. Cancer Cell, 19(1), 17-30.

背景介绍
  • IDH1 (Isocitrate Dehydrogenase 1) is a key metabolic enzyme located in the cytoplasm and peroxisomes. Its primary physiological function is to catalyze the oxidative decarboxylation of isocitrate to produce alpha-ketoglutarate (α-KG), while concurrently reducing NADP+ to NADPH. This reaction is crucial for cellular metabolism, as it links the tricarboxylic acid (TCA) cycle with lipid metabolism and provides NADPH, a major reducing agent essential for antioxidant defense and anabolic biosynthesis.A seminal discovery in cancer biology revealed that specific heterozygous somatic mutations in the IDH1 gene, most commonly at the R132 residue, are frequent drivers in several cancers. These include gliomas (especially secondary glioblastoma and oligodendroglioma), acute myeloid leukemia (AML), cholangiocarcinoma, and chondrosarcomas. The mutant enzyme acquires a neomorphic activity: instead of producing α-KG, it reduces α-KG to D-2-hydroxyglutarate (D-2HG). This oncometabolite, D-2HG, accumulates to high levels and functions as a competitive inhibitor of multiple α-KG-dependent dioxygenases. This inhibition leads to profound epigenetic dysregulation (via inhibition of histone and DNA demethylases) and blockade of cellular differentiation, thereby promoting tumorigenesis.
操作步骤

  • 实验方法

    实验原理：检测IDH1蛋白氧化脱羧异柠檬酸生成2-酮戊二酸的能力。

    实验材料

    1. 检测缓冲液：25 mM Tris, 0.5 mM MnCl2, 5 mM DTT, pH 7.5

    2. IDH1 His Tag Protein, Human

    3. 底物1：DL-Isocitricacid (MCE, Catalog # HY-W009362)

    4. 底物2：NADP+ (Aladdin, Catalog # N101669)

    5. 96 Well Clear Plate (BIOFIL, Catalog#011096)

    6. 读板器（PerkinElmer，ABS, 340 nm, 动力学模式, 60s/cycle,10 cycles）

    实验步骤

    1. 在检测缓冲液中将IDH1蛋白稀释至0.8 µg/mL、0.4 µg/mL。

    2. 制备底物混合物：在检测缓冲液中分别将NADP+和异柠檬酸稀释至1 mM和2 mM。

    3. 向黑色孔板中加入50 µL稀释后的IDH1蛋白（步骤1），然后通过加入50 µL混合底物来启动反应。同时需设置含50 µL的检测缓冲液和50 µL混合底物的空白对照组。

    4. 在340 nm波长下（底部读数）以动力学模式读取孔板10分钟。

    5. 计算特异性活性：

    Specific Activity (pmol/min/µg) = Slope (OD/min) x well volume (L) x 1012pmol/mol

    ext. coeff (M-1cm-1) x path corr. (cm) x amount of enzyme (μg)

    斜率：经底物空白校正

    消光系数：使用消光系数 6270 M⁻¹cm⁻¹

    光程校正：使用光程校正 0.320 cm

  • 电泳

    • 2μg (R: reducing condition, N: non-reducing condition).

  • 体积排阻色谱(SEC-HPLC)

    • The purity of IDH1 His Tag Protein, Human is more than 90% determined by SEC-HPLC.

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