Active Recombinant PNGase F

• Cat.No.: PNGase F-15
• Product Overview: PNGase F is recombinantly expressed in E.coli. Residual Host Cell Protein: <50 ppm

Specification

• Source: E.coli
• Tag: Non
• Description: It can selectively hydrolyze all N-glycan chains linked to Asn in proteins, except those containing α1-3 fucosidic bonds. The cleavage site is the glycosidic bond between Asn and the innermost GlcNAc. This causes the glycoprotein to release the Asn-linked sugar molecules and converts Asn to Asp. The cleaved glycan types include high-mannose, hybrid, and complex oligosaccharides.
• Form: Lyophilized
• Purity: ≥ 95% by SDS-PAGE
• Advantages: 1. High purity: It is free from contamination by other proteases and has no other endo-and exo-glycosidase activities, with a purity of ≥ 95%; 2. High stability: Each batch of PNGase F undergoes strict quality control to ensure batch-to-batch stability of the product; 3. High specific activity: It can effectively and completely release N-linked glycans; 4. Compatible with HPLC/ MS: The lyophilized form of PNGase F does not contain glycerol, which helps to achieve optimal results in HPLC and MS analyses; 5. This product can cleave glycoproteins under denaturing and non-denaturing conditions.
• Protein Content: ≥ 20 KU
• Unit definition: One unit is defined as the amount of the enzyme required to remove more than 95% of the carbohydrates from 10 μg of denatured RNase B in a 10 μL reaction system at 37 centigrade for 1 hour.
• Usage: 1. Protein Reconstitution Centrifuge the centrifuge tube containing lyophilized PNGase F at 100-200 g for 2 minutes. Carefully open the cap and add pure water to achieve a final concentration of 500 U/μL. Invert the tube up and down to mix thoroughly as much as possible. After mixing, centrifuge at 100-200 g for 2 minutes. The reconstituted PNGase F should be stored at-20 centigrade for 6 months, and users should try to avoid repeated freeze-thaw cycles. 2. Denaturing enzymatic digestion Dissolve 1-20 μg of glycoprotein in deionized water. Add 1 μL of 10× glycoprotein denaturation buffer and adjust the volume to 10 μL with deionized water. Incubate at 100 centigrade for 10 min. Transfer to ice and centrifuge for 10 s. Add 2 μL of 10× PNGase F digestion reaction buffer, 2 μL of 10% NP-40, and 5 μL of deionized water. Mix well. Add 1 μL of the reconstituted PNGase F. Incubate at 37 centigrade for 60 min. Analyze the effect of enzymatic digestion. 3. Non-denaturing enzymatic digestion Dissolve 1-20 μg of glycoprotein in deionized water. Add 1 μL of 10× PNGase F digestion reaction buffer and 2-4 μL of the reconstituted PNGase F, then adjust the volume to 10 μL with deionized water. Incubate at 37 centigrade for 2 hours to overnight. Analyze the effect of enzymatic digestion.
• Note: 1. Denaturing enzymatic digestion has a relatively high efficiency and should be attempted first. If denaturing enzymatic digestion cannot be carried out (for example, precipitation occurs during the denaturation process), non-denaturing enzymatic digestion can be tried. However, the digestion efficiency may decrease. It is recommended to increase the amount of enzyme and extend the digestion time; 2. After reconstitution, try to avoid repeated freeze-thaw cycles for this product; 3. Please wear lab coat and disposable gloves when using; 4. This product should not be used directly for clinical diagnosis and treatment.
• Storage: The PNGase F, Lyophilized should be stored at 2-8 centigrade. The 10× glycoprotein denaturation buffer, 10× PNGase F digestion reaction buffer and 10% NP-40 should be stored at-20 centigrade and can be preserved for at least 12 months.
• Shipping: Dry ice or ice packs
• Reconstitution: Ultrapure water

Gene Information

• Official Symbol: PNGase F