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Recombinant SUMO Protease, Yeast

Recombinant SUMO Protease, Yeast

货号: UA070004
价格: 830
规格: 1kU
介绍: -
其他: -
产品规格

  • 物种

    Yeast

  • 表达宿主

    E.coli

  • 分子量

    27 kDa (Reducing)

  • 纯度

    ＞95% by SDS-PAGE and RP-HPLC

  • 内毒素含量

    <1EU/μg

  • 活性

    10U/μl

  • 标记

    Unconjugated

  • 标签

    No Tag

  • 性状

    Liquid

  • 缓冲体系

    20mM PB, pH7.4, 200mM NaCl, 0.1% NP-40, 0.5mM DTT, 50%(v/v) Glycerol

  • 储存条件

    · 12 months from date of receipt, -20 to -70 °C as supplied. 
    · 6 months, -20 to -70 °C under sterile conditions after reconstitution.
    · 1 week, 2 to 8 °C under sterile conditions after reconstitution.  
    · Please avoid repeated freeze-thaw cycles.

  • 稀释度

背景介绍

  • Saccharomyces cerevisiae-derived Small ubiquitin-like modifier (SUMO, Smt3) is commonly used as a protein fusion domain to facilitate expression and purification of recombinant proteins, and a Saccharomyces cerevisiae-derived SUMO-specific protease(Ulp1) is then used to remove SUMO tag from these proteins in a ‘scarless’ manner. SUMO Protease cleaves in a highly specific manner, recognizing the tertiary structure of the SUMO tag, rather than an amino acid sequence, and hydrolyzes the peptide bond in the x-Gly-Gly-x sequence after the Gly-Gly bond at the C-terminus of the SUMO tag. The SUMO Protease cleavage proteins over wide ranges of temperature (4℃-30℃), ionic strengths(0-400 mM NaCl) and pH(7.0-9.0), and easily removed from the cleavage reaction by Immobilized Metal Affinity chromatography (IMAC).

产品组分

  • 1. SUMO Protease;

    2. 10X SUMO Protease Buffer + Salt: 500 mM PB, pH 7.4, 2% Igepal (NP-40), 1.5 M NaCl, 10 mM DTT;

    3. 10X SUMO Protease Buffer – Salt: 500 mM PB, pH 7.4, 2% Igepal (NP-40), 10 mM DTT;

操作步骤

  • 1.Add the following to a microcentrifuge tube:

    Fusion Protein

    20μg

    10X SUMO Protease Buffer +/– Salt

    5μl

    SUMO  Protease

    10U

    ddH2O

    To 50μl

    2. Mix and incubate at 30°C, Remove 5μl aliquots at 1, 2, 4, and 6 hours.

    3. Analyze by SDS-PAGE. 

    • Keep the concentration of Imidazole less then 150mM, or the activity of the SUMO Protease can be adversely affected.
    • For most fusion proteins, SUMO Protease functions optimally in a reaction mixture containing 150 mM NaCl; however, conditions may be optimized by varying the NaCl concentration from 100 mM to 300 mM. Remember to take into account the contribution of salt from the enzyme and from your substrate. When setting up your cleavage reaction, use the appropriate 10X SUMO Protease Buffer +/- Salt.
    • Researchers need to optimize their specific reaction conditions. As an initial suggestion, 20 units of SUMO protease can be used per 40μg of target protein for 1 hour at 30 °C, or overnight at 2–8 °C. The cleavage efficiency can then be estimated by SDS-PAGE.
酶活定义
  • One unit of SUMO Protease cleaves ≥85% of 2 μg control substrate in 1 h at 30°C.

  • 生物活性JSON

    • The substrate of 2 μg fusion protein was digested by SUMO Protease at 30 ℃ for 1 hour. The addition amount of SUMO Protease were 0U, 0.25U, 0.5U, 0.75U and 1U, respectively. The substrate is a fusion protein with a molecular weight of 20 KD and digested completely into two bands.

  • 电泳JSON

    • 1μg (N: non-reducing condition, R: reducing condition).

  • 反相高效液相色谱JSON(RP-HPLC)