PNGase F (with His-tag)

PNGase F (with His-tag)

REF: AS050147-AS050148

Storage and transportation conditions

-20℃

Product composition

Product introduction

PNGase F(Peptide N-glycosidase F) is derived from ElizabethkingiamiricolaIt is a highly efficient amide hydrolase that cleaves high-mannose, heterozygous, and complex oligosaccharide proteins linked by asparagine. PNGase F The cutting check point is the inside of the glycoprotein The amide bond between N-acetylglucosamine (GlcNAc) and aspartamide residues converts aspartamide to aspartic acid after enzymatic hydrolysis.

This product is recombinant expressed by Escherichia coli with His Label.  contain 50% glycerol is often used for the deglycosylation of antibodies and related proteins.

Definition of enzyme activity unit

An enzyme activity unit (U) refers to the amount of enzyme required to remove more than 95% carbohydrates from 10 μg of denatured RNase B for 1 h at 37 ° C in a 10 μl reaction system.

inactivation condition

Incubate at 75 ℃ for 10 min.

Quality Control

Protein purity detection

use SDS-PAGE Gel electrophoresis detection, the protein purity is not less than 95%。

Determination of glycosidase and protease activities

No detectable endoglycosidase F1、F2 or F3 activity; no detectable protease activity.

How to use

1. Glycosylation under denaturing conditions

▲ 1 ~ 20 μg Glycoprotein,   1 μl  10× Denaturing Buffer and Mix ddH2O (if necessary) to a total volume of 10 μl.

Note: 10 × Denaturing Buffer may appear white precipitate when stored at low temperature, and it can be incubated at 37 ° C to dissolve before use.

② After the reaction at 100 ° C for 10 min to denature the glycoprotein, cool on ice and centrifuge for 10 s;

③ SMD 2 μl 10 × PNGase F Bucated er, 2 μl 10% NP-40, ddH2O Make the total reaction volume 20 μl

④ Add 1-2 μl of PNGase F, mix gently, and incubate at 37 ° C for 1-3 h.

2. Glycosylation under non-denaturing conditions

① 将 1~20 μg 糖蛋白，  2  μl  10× PNGase F Buffer and ddH2O ( It is necessary to mix to the total volume 20 μl

② Add 2~ 5 μl PNGase F, mix gently;

③ Incubate at 37 ° C for 4 to 24 hours.

PNGase F removal of

This product comes with His label, after the reaction, you can choose a different label from the target glycoprotein, By affinity chromatography PNGase F was removed.