PNGase F is the most effective enzymatic method for removing almost all N-linked oligosaccharides from glycoproteins. PNGase F is an amidase, which cleaves between the innermost GlcNAc and asparagine residues of high mannose, hybrid, and complex oligosaccharides.
Leaves N-glycan core oligosaccharides intact and suitable for further analysis
Non-recombinant with no detectable endoglycosidase F1, F2 or F3 contamination
≥ 95% purity, as determined by SDS-PAGE and intact ESI-MS
Stored in 50% glycerol
Optimal activity and stability for up to 24 months
Peptide -N-Glycosidase F, also known as PNGase F, is an amidase that cleaves between the innermost GlcNAc and asparagine residues of high mannose, hybrid, and complex oligosaccharides from N-linked glycoproteins (1)
Detailed Specificity:PNGase F is not able to cleave N-linked glycans from glycoproteins when the innermost GlcNAc residue is linked to an α1-3 Fucose residue. This modification is most commonly found in plant and some insect glycoproteins.
Product Source
PNGase F is purified from Flavobacterium meningosepticum (3) and it is free of proteases and Endo F activities.
Glycosidase Recognition Site
This product is related to the following categories:
The following reagents are supplied with this product:
NEB #
Component Name
Component #
Stored at (°C)
Amount
Concentration
P0704S
-20
PNGase F
P0704SVIAL
-20
1 x 0.03 ml
500,000 units/ml
GlycoBuffer 2
B3704SVIAL
-20
1 x 1 ml
10 X
Glycoprotein Denaturing Buffer
B1704SVIAL
-20
1 x 1 ml
10 X
NP-40
B2704SVIAL
-20
1 x 1 ml
10 %
P0704L
-20
PNGase F
P0704LVIAL
-20
1 x 0.15 ml
500,000 units/ml
GlycoBuffer 2
B3704SVIAL
-20
1 x 1 ml
10 X
Glycoprotein Denaturing Buffer
B1704SVIAL
-20
1 x 1 ml
10 X
NP-40
B2704SVIAL
-20
1 x 1 ml
10 %
Properties & Usage
Unit Definition
One unit is defined as the amount of enzyme required to remove > 95% of the carbohydrate from 10 µg of denatured RNase B in 1 hour at 37°C in a total reaction volume of 10 µl.
Unit Definition Assay: 10 µg of RNase B are denatured with 1X Glycoprotein Denaturing Buffer (0.5% SDS, 40 mM DTT) at 100°C for 10 minutes. After the addition of NP-40 and GlycoBuffer 2, two-fold dilutions of PNGase F are added and the reaction mix is incubated for 1 hour at 37°C. Separation of reaction products are visualized by SDS-PAGE.
1X Glycoprotein Denaturing Buffer 0.5% SDS 40 mM DTT
Since PNGase F activity is inhibited by SDS, it is essential to have NP-40 present in the reaction mixture. Why this non-ionic detergent counteracts the SDS inhibition is unknown at present.
To deglycosylate a native glycoprotein, longer incubation time as well as more enzyme may be required.
PNGase F will not cleave N-linked glycans containing core α1-3 Fucose.
Typical reaction conditions: Please see Protocols
References
Maley, F. et al. (1989). Anal. Biochem. 180, 195-204.
Tretter, V. et al. (1991). Eur. J. Biochem.. 199, 647-652.
Plummer, T.H. Jr. and Tarentino, A.L. (1991). Glycobiology. 1, 257-263.
PNGase F
You can use this enzyme under native or denaturing conditions
Under native conditions we recommend adding more enzyme and using longer incubation times
PNGase F activity is inhibited by SDS, therefore under denaturing conditions it is essential to have NP-40 present in the reaction mixture in a 1:1 ratio.
PNGase F will not cleave N-linked glycans containing core a1-3 Fucose (PNGase A must be used in this instance)
Enzyme activity varies at different temperatures: 37°C - 100%; 30°C - 100%; 23°C - 65%; 17°C - 40% and 3°C - 0%
A good positive control substrate is RNase B
Liu H., Zou X., Li T., Wang X., Yuan W., Chen Y., Han W. (2015) Enhanced production of secretory glycoprotein VSTM1-v2 with mouse IgGκ signal peptide in optimized HEK293F transient transfection Sci Rep; PubMedID: 26140918, DOI: 10.1038/srep11603
Yang W., Zhang Y., Zhou X., Zhang W., Xu X., Chen R., Meng Q., Yuan J., Yang P., Yao B. (2015) Production of a Highly Protease-Resistant Fungal α-Galactosidase in Transgenic Maize Seeds for Simplified Feed Processing Sci Rep; 5, 11603. PubMedID: 26053048, DOI: 10.1038/srep11603
Chen J., Fang M., Zhao YP., Yi CH., Ji J., Cheng C., Wang MM., Gu X., Sun QS., Chen XL., Gao CF. (2015) Serum N-Glycans: A New Diagnostic Biomarker for Light Chain Multiple Myeloma Sci Rep; 5, 11603. PubMedID: 26075387, DOI: 10.1038/srep11603
Netsirisawan P., Chokchaichamnankit D., Srisomsap C., Svasti J., Champattanachai V. (2015) Proteomic Analysis Reveals Aberrant O-GlcNAcylation of Extracellular Proteins from Breast Cancer Cell Secretion Mol Biol Cell; 26, 2168-80. PubMedID: 26136220
Brown EP., Normandin E., Osei-Owusu NY., Mahan AE., Chan YN., Lai JI., Vaccari M., Rao M., Franchini G., Alter G., Ackerman ME. (2015) Microscale purification of antigen-specific antibodies Sci Rep; PubMedID: 26078040, DOI: 10.1038/srep11603
Shakiba N., White C.A., Lipsitz Y.Y., Yachie-Kinoshita A., Tonge P.D., Hussein S.M.I., Puri M.C., Elbaz J., Morrissey-Scoot J., Li M., Munoz J., Benevento M., Rogers I.M., Hanna J.H., Heck A.J.R., Wollscheid B., Nagy A., Zandstra P.W. (2015) CD24 tracks divergent pluripotent states in mouse and human cells. Nat Commun; 6, 7329. PubMedID: 26076835
Malsburg K., Shao S., Hegde RS. (2015) The ribosome quality control pathway can access nascent polypeptides stalled at the Sec61 translocon Mol Biol Cell; 26, 2168-80. PubMedID: 25877867, DOI: 10.1091/mbc.E15-01-0040
Orizio F., Damiati E., Giacopuzzi E., Benaglia G., Pianta S., Schauer R., Schwartz-Albiez R., Borsani G., Bresciani R., Monti E. (2015) Human sialic acid acetyl esterase: Towards a better understanding of a puzzling enzyme. Glycobiology; 25, 992-1006. PubMedID: 26022516
Singh S., Kuntal P., Yadav J., Tang H., Partyka K., Kletter D., Hsueh P., Ensink E., Kc B., Hostetter G., Xu E.H., Bern M., Smith D.F., Mehta A.S., Brand R., Melcher K., Haab B.B. (2015) Upregulation of glycans containing 3' fucose in a subset of pancreatic cancers uncovered using fusion-tagged lectins. J Proteome Res; 14, 2594-605. PubMedID: 25938165
Go E.P., Herschhorn A., Gu C., Castillo-Menendez L., Zhang S., Mao Y., Chen H., Ding H., Wakefield J.K., Hua D., Liao H.X., Kappes J.C., Sodroski J., Desaire H. (2015) Comparative Analysis of the Glycosylation Profiles of Membrane-Anchored HIV-1 Envelope Glycoprotein Trimers and Soluble gp140. J Virol; 89, 8245-57. PubMedID: 26018173
Haramoto Y., Takahashi S., Oshima T., Onuma Y., Ito Y., Asashima M. (2015) Insulin-like factor regulates neural induction through an IGF1 receptor-independent mechanism. Mol Biol Cell; 26, 2168-80. PubMedID: 26112133, DOI: 10.1091/mbc.E15-01-0040
Wang L., Zhang X., Pang N., Xiao L., Li Y., Chen N., Ren M., Deng X., Wu J. (2015) Glycation of vitronectin inhibits VEGF-induced angiogenesis by uncoupling VEGF receptor-2-αvβ3 integrin cross-talk Mol Biol Cell; 26, 2168-80. PubMedID: 26111058, DOI: 10.1091/mbc.E15-01-0040
Wang J., Hilchey SP., Hyrien O., Huertas N., Perry S., Ramanunninair M., Bucher D., Zand MS. (2015) Multi-Dimensional Measurement of Antibody-Mediated Heterosubtypic Immunity to Influenza. Sci Rep; 5, 11603. PubMedID: 26103163, DOI: 10.1038/srep11603
Pritchard L.K., Harvey D.J., Bonomelli C., Crispin M., Doores K.J. (2015) Cell- and Protein-Directed Glycosylation of Native Cleaved HIV-1 Envelope. J Virol; 89, 8932-44. PubMedID: 26085151
Holemans T., Sørensen DM., Veen S., Martin S., Hermans D., Kemmer GC., Haute C., Baekelandt V., Pomorski TG., Agostinis P., Wuytack F., Palmgren M., Eggermont J., Vangheluwe P. (2015) A lipid switch unlocks Parkinson's disease-associated ATP13A2 Proc Natl Acad Sci U S A; 112, 9040-5. PubMedID: 26134396
Asazuma HM., Sohn BH., Kim Y.S., Kuo CW., Khoo KH., Kucharski CA., Fraser MJ., Jarvis DL. (2015) Targeted Glycoengineering Extends the Protein N-glycosylation Pathway in the Silkworm Silk Gland Sci Rep; PubMedID: 26163436, DOI: 10.1038/srep11603
AlSalmi W., Mahalingam M., Ananthaswamy N., Hamlin C., Flores D., Gao G., Rao V.B. (2015) A New Approach to Produce HIV-1 Envelope Trimers: BOTH CLEAVAGE AND PROPER GLYCOSYLATION ARE ESSENTIAL TO GENERATE AUTHENTIC TRIMERS. J Biol Chem; 290, 19780-95. PubMedID: 26088135
Julien M., Chauvet S., Scheckenbach KE., Alfaidy N., Chanson M., Benharouga M. (2015) Involvement of the heterodimeric interface region of the nucleotide binding domain-2 (NBD2) in the CFTR quaternary structure and membrane stability Sci Rep; PubMedID: 26083625, DOI: 10.1038/srep11603
Beata O., Jarząb A., Kratz E., Zimmer M., Gamian A., Ferens-Sieczkowska M. (2015) Terminal Mannose Residues in Seminal Plasma Glycoproteins of Infertile Men Compared to Fertile Donors Int J Mol Sci; 16, 14933-50. PubMedID: 26147424, DOI: 10.3390/ijms160714933
Noble GP., Wang DW., Walsh DJ., Barone JR., Miller MB., Nishina KA., Li S., Supattapone S. (2015) A Structural and Functional Comparison Between Infectious and Non-Infectious Autocatalytic Recombinant PrP Conformers Sci Rep; 5, 11603. PubMedID: 26125623, DOI: 10.1038/srep11603
Itahana Y, Han R, Barbier S, Lei Z, Rozen S, Itahana K (2014) The uric acid transporter SLC2A9 is a direct target gene of the tumor suppressor p53 contributing to antioxidant defense Oncogene; PubMedID: 24858040, DOI: 10.1038/onc.2014.119
Rosenbaek LL, Kortenoeven ML, Aroankins TS, Fenton RA (2014) Phosphorylation decreases ubiquitylation of the thiazide-sensitive cotransporter NCC and subsequent clathrin-mediated endocytosis J Biol Chem; 289(19), 13347-61. PubMedID: 24668812, DOI: 10.1074/jbc.M113.543710
Botto L, Cunati D, Coco S, Sesana S, Bulbarelli A, Biasini E, Colombo L, Negro A, Chiesa R, Masserini M, Palestini P (2014) Role of lipid rafts and GM1 in the segregation and processing of prion protein PLoS One; 9(5), e98344. PubMedID: 24859148, DOI: 10.1371/journal.pone.0098344
Wright CR, Brown EL, Della-Gatta PA, Ward AC, Lynch GS, Russell AP (2014) G-CSF does not influence C2C12 myogenesis despite receptor expression in healthy and dystrophic skeletal muscle Front Physiol; 5, 170. PubMedID: 24822049, DOI: 10.3389/fphys.2014.00170
Wicht O, Burkard C, de Haan CA, van Kuppeveld FJ, Rottier PJ, Bosch BJ (2014) Identification and Characterization of a Proteolytically Primed Form of the Murine Coronavirus Spike Proteins after Fusion with the Target Cell J Virol; 88(9), 4943-52. PubMedID: 24554652, DOI: 10.1128/JVI.03451-13
Kwon HM, Lee KH, Han BW, Han MR, Kim DH, Kim DE (2014) An RNA aptamer that specifically binds to the glycosylated hemagglutinin of avian influenza virus and suppresses viral infection in cells PLoS One; 9(5), e97574. PubMedID: 24835440, DOI: 10.1371/journal.pone.0097574
Stech M, Quast RB, Sachse R, Schulze C, Wüstenhagen DA, Kubick S (2014) A continuous-exchange cell-free protein synthesis system based on extracts from cultured insect cells PLoS One; 9(5), e96635. PubMedID: 24804975, DOI: 10.1371/journal.pone.0096635
Haller G, Li P, Esch C, Hsu S, Goate AM, Steinbach JH (2014) Functional characterization improves associations between rare non-synonymous variants in CHRNB4 and smoking behavior PLoS One; 9(5), e96753. PubMedID: 24804708, DOI: 10.1371/journal.pone.0096753
Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.
Specifications & Change Notifications
The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]
The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]
Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.
This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.
New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.
Based on your Freezer Program type, you are trying to add a product to your cart that is either not allowed or not allowed with the existing contents of your cart. Please review and update your order accordingly If you have any questions, please contact Customer Support at freezers@neb.com or 1-800-632-5227 x 8.
You have been idle for more than 20 minutes, for your security you have been logged out. Please sign back in to continue your session.
Institution Changed
Your profile has been mapped to an Institution, please sign back for your profile updates to be completed.
Sign in to your NEB account
To save your cart and view previous orders, sign in to your NEB account. Adding products to your cart without being signed in will result in a loss of your cart when you do sign in or leave the site.
Powered by Bioz
