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Remove-iT® PNGase F |NEB酶试剂 New England Biolabs

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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

Remove-iT® PNGase F |
 
Remove-iT® PNGase F is an amidase which cleaves between the innermost GlcNAc and asparagine residues of high mannose, hybrid, and complex oligosaccharides from N-linked glycoproteins (1). Remove-iT PNGase F is tagged with a chitin binding domain (CBD) for easy removal from a reaction and is supplied glycerol free for optimal performance in HPLC and MS intensive methods.

产品来源

Remove-iT PNGase F is purified from Elizabethkingia miricola (formerly Flavobacterium meningosepticum).

特异性

Remove-iT® PNGase F |
产品类别:
Endoglycosidases Products,
Proteome Analysis Products

应用:
Expression Systems,
Glycan Sequencing,
Proteomics,

Recombinant Glycoprotein Expression,

Glycoprotein Analysis

  • 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
    • P0706S     4    
        Remove-iT® PNGase F P0706SVIAL 4 1 x 0.03 ml 225,000 units/ml
        GlycoBuffer 2 B3704SVIAL -20 1 x 1 ml 10 X
        DTT B0706SVIAL -20 1 x 1 ml 0.4 M
    • P0706L     4    
        Remove-iT® PNGase F P0706LVIAL 4 1 x 0.15 ml 225,000 units/ml
        GlycoBuffer 2 B3704SVIAL -20 1 x 1 ml 10 X
        DTT B0706SVIAL -20 1 x 1 ml 0.4 M

  • 特性和用法

    单位定义

    One unit is defined as the amount of enzyme required to remove > 95% of the carbohydrate from 5 μg of DTT denatured RNase B in 1 hour at 37°C in a total reaction volume of 10 μl.

    反应条件

    1X GlycoBuffer 2
    Incubate at 37°C

    1X GlycoBuffer 2
    50 mM Sodium Phosphate
    (pH 7.5 @ 25°C)

    贮存溶液

    20 mM Tris-HCl
    50 mM NaCl
    5 mM EDTA
    pH 7.5 @ 25°C

    热失活

    75°C for 10 minutes

    分子量

    实际: 41000 daltons

    单位活性检测条件

    5 µg of RNase B are denatured with 1X DTT at 55°C for 10 minutes. After the addition of 1X GlycoBuffer 2, two-fold dilutions of Remove-iT PNGase F are added and the reaction mix is incubated for 1 hour at 37°C. Separation of reaction products is visualized by SDS-PAGE.

  • 相关产品

    相关产品

    • RNase B（对照底物）
    • 几丁质磁珠
    • 6 孔磁性分离架
    • 12 孔磁性分离架
    • 胎球蛋白
    • Endo S
    • 糖苷内切酶 D
    • p0704-pngase-f

  • 注意事项

    1. To deglycosylate a native glycoprotein, longerincubation time, as well as more enzyme, maybe required.
    2. Using typical RNase B denaturing conditions with NEB Glycoprotein Denaturing Buffer,containing SDS and DTT, Remove-iT PNGase F yields a higher concentration of 500,000 U/ml.
    3. If using Remove-iT PNGase F under typical PNGase F denaturing conditions, it is essential to have NP-40 in the reaction mixture as Remove-iT PNGase F is inhibited by SDS. It is not known why this non-ionic detergent counteracts the SDS inhibition.
    4. Remove-iT PNGase F will not cleave N-linkedglycans containing core α1-3 Fucose.
    5. Recommended storage temperature is 4°C,avoid repeat freeze-thaw cycles
    6. Removal of Remove-iT PNGase F from the deglycosylation reaction can be scaled up linearly with larger volumes of chitin magnetic beads.
    7. Chitin Magnetic Beads Binding Capacity is0.4 µg/µl of CBD-tagged protein.
    8. To remove 1-5 µl of Remove-iT PNGase F use 50 µl of chitin magnetic beads.

  • 参考文献

    1. Maley, F. et al. (1989). Anal. Biochem. 180, 195-204.

操作说明、说明书 & 用法

  • 操作说明

    1. Remove-iT® PNGase F Magnetic Chitin Bead Protocol (P0706)
    2. Reaction Conditions for Remove-iT® PNGase F (P0706)

  • 应用实例

    • AppNote_Remove-iT_PNGase_F_Effective_Release_and_Recovery_of_Neutral_and_Sialylated_N-glycans
    • AppNote_Proteomics_Fast_and_Efficient_Antibody_Deglycosylation_using_Rapid_PNGase_F
    • AppNote_Glycan_Analysis_of_Murine_IgG2a_by_Enzymatic_Digestion_with_PNGase_F_and_Trypsin
    • Glycan Analysis of Murine IgG2a by Enzymatic Digestion with PNGase F and Trypsin, Followed by Mass Spectrometric Analysis
    • Remove iT PNGase F Effective Release and Recovery of Neutral and Sialylated N glycans

工具 & 资源

  • 选择指南

    • Endoglycosidase Selection Chart

FAQs & 问题解决指南

  • FAQs

    1. What is the tag on PNGase F?
    2. I tried deglycosylating my glycoprotein with Remove-iT® PNGase F but did not see removal of the carbohydrate. What could be the problem?
    3. What is the difference between PNGase F and Remove-iT® PNGase F?
    4. What is the difference between Remove-iT® PNGase F and Endo H?
    5. How much Remove-iT® PNGase F should I use to remove my carbohydrate under native or DTT denaturing conditions?
    6. Does Remove-iT® PNGase F work in Urea?
    7. What are the typical reaction conditions for Remove-iT® PNGase F?
    8. How do I eliminate Remove-iT® PNGase F from a reaction?
    9. What is the binding capacity of the Magnetic Chitin Beads used to eliminate Remove-iT® PNGase F?
    10. Is Remove-iT® PNGase F compatible with downstream analysis such as HPLC and Mass Spectrometry?
    11. Why have the NEB Glycosidase enzymes changed reaction buffers? What are the new reaction buffers and can I still use an enzyme with its old buffer? Where can I find the composition of the old buffers?
    12. What is a good endoglycosidase substrate?
    13. Do detergents inhibit exoglycosidases/endoglycosidases?
    14. What are Glycosidases and their uses?