标签归档：phosphatase

Lambda Protein Phosphatase (Lambda PP) |NEB酶试剂 New England Biolabs

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产品信息

Lambda Protein Phosphatase (Lambda PP) is a Mn2+-dependent protein phosphatase with activity towards phosphorylated serine, threonine and tyrosine residues. It is the 221 amino-acid product of the ORF221 open reading frame on bacteriophage lambda (1,2).

重点

  • Dual Specificity: protein serine/threonine/tyrosine phosphatase 
  • Bacteriophage lambda, recombinant (E. coli)

    • 产品来源

    Isolated from a strain of E. coli that carries the bacteriophage lambda ORF221 open reading frame under the control of a T7 expression system.

    产品类别:
    Protein Phosphatases Products

    应用:
    Protein Phosphatases and Kinases

    • 产品组分信息

      本产品提供以下试剂或组分：

      NEB # 名称 组分货号 储存温度 数量 浓度
      • P0753S     -80    
          Lambda Protein Phosphatase (Lambda PP) P0753SVIAL -80 1 x 0.05 ml 400,000 units/ml
          NEBuffer Pack for Protein MetalloPhosphatases (PMP) B0761SVIAL -20 1 x 1 ml 10 X
          MnCl2 B1761SVIAL -20 1 x 1 ml 10 X
      • P0753L     -80    
          Lambda Protein Phosphatase (Lambda PP) P0753LVIAL -80 1 x 0.25 ml 400,000 units/ml
          NEBuffer Pack for Protein MetalloPhosphatases (PMP) B0761SVIAL -20 1 x 1 ml 10 X
          MnCl2 B1761SVIAL -20 1 x 1 ml 10 X

    • 特性和用法

      单位定义

      One unit is defined as the amount of enzyme that hydrolyzes 1 nmol of p-Nitrophenyl Phosphate (50 mM) (NEB #P0757) in 1 minute at 30°C in a total reaction volume of 50 µl.

      反应条件

      1X NEBuffer Pack for Protein MetalloPhosphatases (PMP)
      Supplement with 1 mM MnCl2
      Incubate at 30°C

      1X NEBuffer Pack for Protein MetalloPhosphatases (PMP)
      50 mM HEPES
      100 mM NaCl
      2 mM DTT
      0.01% Brij 35
      (pH 7.5 @ 25°C)

      贮存溶液

      50 mM HEPES
      100 mM NaCl
      2 mM DTT
      0.01% Brij 35
      0.1 mM EGTA
      0.1 mM MnCl2
      50% Glycerol
      pH 7.5 @ 25°C

      热失活

      65°C for 60 minutes

      分子量

      理论上的: 25 kDa

      比活度

      800,000 units/mg

      存储注意事项

      • Avoid repeated freeze/thaw cycles.

      货运单

      • Ships on dry ice

    • 优势和特性

      应用特性

      • Lambda PP can be used to release phosphate groups from phosphorylated serine, threonine and tyrosine residues in proteins. Note that different proteins are dephosphorylated at different rates.

    • 相关产品

      相关产品

      • p-Nitrophenyl Phosphate (PNPP)
      • Sodium Orthovanadate (Vanadate)

      单独销售的组分

      • NEBuffer Pack for Protein MetalloPhosphatases (PMP)

    • 注意事项

      1. The following information can be used as suggested initial conditions for dephosphorylation of proteins with Lambda PP.
      2. 100 units of Lambda PP remove ~100% of phosphates (0.5 nmol) in phosphorylated myelin basic protein (phospho-MyBP, 18.5 kDa) in 30 minutes in a 50 µl reaction. The concentration of phospho-MyBP is 10 µM with respect to phosphate.
      3. The Protein Serine/threonine Phosphatase (PSP) activity of Lambda PP is assessed on MyBP phosphorylated exclusively on serine/threonine residues with cAMP-dependent Protein Kinase. The Protein Tyrosine Phosphatase (PTP) activity is assessed on MyBP phosphorylated exclusively on tyrosine residues with Abl Protein Tyrosine Kinase.
      4. Lambda PP is active on phosphorylated histidine residues (2).
      5. 0ptimal incubation times and enzyme concentrations must be determined empirically for each particular substrate. 
      6. If the source of phosphorylated protein is a crude extract of cells or tissue, it is very important to include the appropriate protease inhibitors in the lysis buffer and to use shorter incubation time for dephosphorylation.
      7. The following levels of inhibition of Lambda PP (100 units) are found when the reaction buffer and MnCl2 are supplemented with: 
        • 10 mM Sodium Orthovanadate (3): 80% 
        • 10 mM Sodium Orthovanadate, 50 mM Sodium Fluoride: 90%
        • 50 mM EDTA: 95% 
        • 1% Triton X-100: no 
        • 0.4% Nonidet P-40: no 
        • 0.025% Tween 20: no 
        • 0.5 M NaCl: 5% 
        • ATP Mix (10 mM MgCl2, 0.1 mM ATP): no
        • Protease Inhibitor Cocktail*: 10% 

        *Pepstatin A, leupeptin and aprotinin, 10 µg/ml each, 0.5 mM PMSF and 1 mM benzamidine

    • 参考文献

      1. Cohen, P.T.W. and Cohen, P. (1989). Biochem. J.. 260, 931-934.
      2. Zhuo, S. et al. (1993). J. Biol. Chem.. 268, 17754-17761.
      3. Gordon, J.A. (1991). Methods Enzymol.. 201, 477-482.

    操作说明、说明书 & 用法

    • 操作说明

      1. Typical reaction protocol for lambda protein phosphatase

    FAQs & 问题解决指南

    • FAQs

      1. Can you recommend a cell lysis buffer that is compatible with Lambda Protein Phosphatase (NEB #P0753)?

    Alkaline Phosphatase Staining Kit

    发表于由

    Alkaline Phosphatase Staining Kit

    品牌：Cosmo Bio
    CAS No.：
    储存条件：4℃ DNF
    纯度：–
    产品编号

    (生产商编号)

    		 等级 规格 运输包装 零售价(RMB) 库存情况 参考值

    PMC-AK20

    		 – 1 Kit – 咨询 – –


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    T4 Polynucleotide Kinase (3′ phosphatase minus) |NEB酶试剂 New England Biolabs

    发表于由

    上海金畔生物科技有限公司代理New England Biolabs（NEB）酶试剂全线产品，欢迎访问官网了解更多产品信息和订购。

    产品信息

    T4 Polynucleotide Kinase catalyzes the transfer and exchange of Pi from the γ position of ATP to the 5´ -hydroxyl terminus of double- and single-stranded DNA and RNA, as well as nucleoside 3´-monophosphates (1-5). This modified version exhibits full kinase activity with no 3´ phosphatase activity (6,7).

    产品来源

    An E. coli strain that carries a plasmid encoding the modified T4 Polynucleotide Kinase gene.

    产品类别:
    Kinases Products,
    RNA Modification

    应用:
    Phosphorylation (Kinase)

    • 产品组分信息

      本产品提供以下试剂或组分：

      NEB # 名称 组分货号 储存温度 数量 浓度
      • M0236S     -20    
          T4 Polynucleotide Kinase (3′ phosphatase minus) M0236SVIAL -20 1 x 0.02 ml 10,000 units/ml
          T4 Polynucleotide Kinase Reaction Buffer B0201SVIAL -20 1 x 1 ml 10 X
      • M0236L     -20    
          T4 Polynucleotide Kinase (3′ phosphatase minus) M0236LVIAL -20 1 x 0.1 ml 10,000 units/ml
          T4 Polynucleotide Kinase Reaction Buffer B0201SVIAL -20 1 x 1 ml 10 X

    • 特性和用法

      单位定义

      One unit is defined as the amount of enzyme catalyzing the incorporation of 1 nmol of acid-insoluble [32P] in 30 minutes at 37°C (1). 

      Unit Assay Conditions: 
      1X T4 Polynucleotide Kinase Reaction Buffer, 66 µM [γ-32P] ATP (5 x 106 cpm/µmol), 0.26 mM 5´-hydroxyl-terminated salmon sperm DNA.

      反应条件

      1X T4 Polynucleotide Kinase Reaction Buffer
      Incubate at 37°C

      1X T4 Polynucleotide Kinase Reaction Buffer
      70 mM Tris-HCl
      10 mM MgCl2
      5 mM DTT
      (pH 7.6 @ 25°C)

      使用浓度

      10,000 units/ml

      贮存溶液

      10 mM Tris-HCl
      50 mM KCl
      1 mM DTT
      0.1 mM EDTA
      50% Glycerol
      0.1 µM ATP
      pH 7.4 @ 25°C

      热失活

      65°C for 20 minutes

      单位活性检测条件

      1X T4 Polynucleotide Kinase Reaction Buffer, 66 µM [γ-32P] ATP (5 x 106 cpm/µmol), 0.26 mM 5´-hydroxyl-terminated salmon sperm DNA.

    • 优势和特性

      Features

      5´ phosphorylation of DNA/RNA for subsequent ligation
      End-labeling of DNA or RNA (2)
      5´ phosphorylation of 3´ phosphorylated mononucleotides to generate a substrate (pNp) that can be added to the 3´ end of DNA or RNA by ligase activity (8)
      5´ end labeling of 3´ phosphorylated oligos (9)

    • 相关产品

      相关产品

      • t1010-monarch-plasmid-miniprep-kit
      • Monarch® DNA 胶回收试剂盒
      • Monarch® PCR & DNA 纯化试剂盒（5 μg）

      单独销售的组分

      • T4 多聚核苷酸激酶反应缓冲液

    • 注意事项

      1. Gaps can be phosphorylated with elevated levels of ATP. Nicks are not phosphorylated efficiently. CTP, GTP, TTP, UTP, dATP or dTTP can be substituted for ATP as a phosphate donor (1).
      2. Dephosphorylation prior to end-labeling can be avoided by utilizing the exchange reaction, although this results in lower specific activity labeling.
      3. Sufficient incorporation levels can be attained using the supplied buffer supplemented with 100 µM ADP in the final reaction. However, higher levels of incorporation with the exchange reaction are achieved when using the buffer described in (2).

    • 参考文献

      1. Richardson, CC. (1981). The Enzymes. 14, 299-314.
      2. Sambrook, J. et al. (1989). Molecular Cloning: A Laboratory Manual. 10.59-10.67,11.31-11.33.
      3. Cameron, V. and Uhlenbeck, O.C. (1977). Biochemistry . 16, 5120-5126.
      4. Berkner, K.L. and Folk, W.R. (1977). J. Biol. Chem.. 252, 3176-3184.
      5. Soltis, D.A. and Uhlenbeck, O.C (1982). J. Biol. Chem.. 257, 11332-11339.
      6. Wang, L.K. and Human, S (2001). J. Biol. Chem.. 276, 26868-26874.
      7. Wang, L.K. and Shuman, S. (2002). Nucl. Acids Res.. 30, 1073-1080.
      8. Vance, JR. and Wilson, T.E. (2001). Mol. Cell. Riot.. 21, 7191-7198.
      9. Interthal, H. et al. (2005). J. Biol. Chem.. 280, 36518-36528.

    操作说明、说明书 & 用法

    • 操作说明

      1. Radioactive Labeling with T4 PNK or T4 PNK (3´ phosphatase minus)
      2. Non-radioactive Phosphorylation with T4 PNK or T4 PNK (3´ phosphatase minus)

    • 使用指南

      • Activity of DNA Modifying Enzymes in rCutSmart™ Buffer

    FAQs & 问题解决指南

    • FAQs

      1. Will T4 PNK (3′ phosphate minus) work in rCutSmart Buffer?

    Bacterial Alkaline Phosphatase from E.coli C75 (BAP) 细菌碱性磷酸酶（E.coli C75）

    发表于由

    Bacterial Alkaline Phosphatase from E.coli C75 (BAP)

    细菌碱性磷酸酶（E.coli C75）

    品牌：Nippon Gene
    CAS No.：
    储存条件：-20℃
    纯度：–
    产品编号

    (生产商编号)

    		 等级 规格 运输包装 零售价(RMB) 库存情况 参考值

    316-01593

    		 – 60U×5 – 咨询 – –


    * 干冰运输、大包装及大批量的产品需酌情添加运输费用


    * 零售价、促销产品折扣、运输费用、库存情况、产品及包装规格可能因各种原因有所变动，恕不另行通知，确切详情请联系上海金畔生物科技有限公司。