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Glycosylated Proteins

Glycosylated proteins are a class of biomolecules where carbohydrate groups are covalently attached to the protein backbone.
These complex structures play vital roles in many biological processes, including cell signaling, immune function, and protein folding.
Optimizing experimental protocols for working with glycosylated proteins is crucial for advancing research in this field.
PubCompare.ai is an AI-driven platform that empowers researchers to locate, compare, and identify the most effective and reproducible methods from literature, preprints, and patents.
This tool simplifies the reseach process and enhances experimental results for studies involving glycosylated proteins.

Most cited protocols related to «Glycosylated Proteins»

1
Structural Analysis of NMDA Receptor ATDs
Cited 18 times
GluN1b and GluN2B ATD proteins were expressed as secreted proteins using the insect cells/baculovirus system and purified using metal-chelate chromatography and size-exclusion chromatography. Crystallization was performed in hanging-drop vapor diffusion configuration in a buffer containing 20% PEG3350, 150 mM KNO3 and 50 mM HEPES-NaOH (pH 7.0) and 3.0–3.5 M NaFormate and 0.1M HEPES-NaOH (pH 7.5) for GluN1b ATD and GluN1b/GluN2B ATDs, respectively. Diffraction data sets obtained at 100K were indexed, integrated, and scaled using HKL2000. The GluN1b ATD structure was solved by the single anomalous diffraction (SAD) phasing method using Se-Met incorporated crystals and the GluN1b/GluN2B ATD structures were solved by molecular replacement using coordinates of GluN1b ATD and GluN2B ATD (PDB code: 3JPW)10 (link). Model refinement was conducted using the program Phenix20 (link). Experiments involving analytical ultracentrifugation and isothermal titration calorimetry were conducted using the purified protein samples in the glycosylated form. Ion channel activities of full-length NMDA receptors were measured by whole-cell recording using cRNA injected Xenopus laevis oocytes using a two electrode voltage-clamp configuration.
Karakas E., Simorowski N, & Furukawa H. (2011). Subunit Arrangement and Phenylethanolamine Binding in GluN1/GluN2B NMDA Receptors. Nature, 475(7355), 249-253.
Publication 2011
Baculoviridae Buffers Calorimetry Cells Chromatography Complementary RNA Crystallization Diffusion Gel Chromatography Glycosylated Proteins GRIN2B protein, human HEPES Insecta Ion Channel Metals N-Methyl-D-Aspartate Receptors Oocytes polyethylene glycol 3350 Proteins Titrimetry Ultracentrifugation Xenopus laevis
2
Structural Characterization of NPC1 and Ebola Virus Glycoprotein Complex
Cited 10 times

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Publication 2016
Amniotic Fluid ATP8A2 protein, human Baculoviridae Chromatography, Affinity Cloning Vectors Crystallization Cytokinesis Diffusion DNA, Complementary Endosomes Escherichia coli Gel Chromatography Glycerin Glycosylated Proteins HEK293 Cells his6 tag Inclusion Bodies Metals Molar Mothers Mucins Mutagenesis, Site-Directed Niemann-Pick C1 Protein Niemann-Pick Disease, Type C1 Peptide Hydrolases Polyethylene Glycols Proteins Sf9 Cells Sodium Acetate Trihydrate Sodium Chloride Strains Thermolysin
3
Automated Transfection of Glycosylated Proteins
Cited 9 times
The vector chosen for development of an automated transfection protocol was pHLsec. This vector was built on the pLEXm backbone (Aricescu et al., 2006 (link)) and contains: pBR322 origin of replication giving the high copy number in Escherichia coli needed to obtain the high amounts of plasmid DNA required for transient transfection; ampicillin resistance; a cytomegalovirus enhancer; chick β-actin promotor to give high levels of expression; the rabbit β-globin intron to increase RNA production; and a poly-A signal to increase RNA stability. Two cell lines were chosen for the development of automated transient transfection protocols for glycosylated mammalian proteins: HEK 293T (ATCC CRL-11268), grown in the presence of the N-glycosylation inhibitor, kifunensine (Chang et al., 2007 (link)) added immediately post-transfection at 1 mg/L final concentration, and the N-acetylglucosaminyltransferase I-negative HEK 293S GnTI− cells (Reeves et al., 2002 (link)), which are unable to synthesize complex glycans. These cell lines were chosen due to ease of handling, robust growth rates, excellent transfection efficiency, high capacity for recombinant protein expression and low cost media requirements. Both cell lines are also routinely used for manual transient transfections, thus allowing a direct comparison between automated and manual protocols.
Zhao Y., Bishop B., Clay J.E., Lu W., Jones M., Daenke S., Siebold C., Stuart D.I., Yvonne Jones E, & Radu Aricescu A. (2011). Automation of large scale transient protein expression in mammalian cells. Journal of Structural Biology, 175(2-2), 209-215.
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Publication 2011
Actins alpha-1,3-mannosyl-glycoprotein beta-1,2-N-acetylglucosaminyltransferase I beta-Globins Cell Lines Cells Cloning Vectors Cytomegalovirus Escherichia coli Glycosylated Proteins Introns kifunensine Mammals Plasmids Poly A Polysaccharides Protein Glycosylation Rabbits Recombinant Proteins Replication Origin Transfection Transients Vertebral Column
4
Quantifying Protein Glycosylation Dynamics
Cited 9 times
The following samples were used to detect glycosylation of the indicated proteins in Fig. 1c: 293T whole cell lysate (Sp1), rat hypothalamus crude nuclear pellet (MeCP2), rat brain detergent-soluble fraction (synapsin IIa, Nup62), whole cell lysate from cultured embryonic neurons (CREB), cytosolic fraction from PUGNAc-treated cultured embryonic neurons (OGA), and p75-OGT purified from Sf9 cells. For Fig. 3a, the liver, hippocampus or cerebral cortex was harvested from adult Sprague Dawley rats, and crude nuclear pellets were processed. Animal protocols were approved by the Institutional Animal Care and Use Committee at Caltech, and the procedures were performed in accordance with the Public Health Service Policy on Humane Care and Use of Laboratory Animals. Cell lysates were prepared as described in the Supplementary Methods. Each sample and its corresponding negative control (lacking ketogalactose probe 1 incorporation) was subjected to chemoenzymatic labeling with PEG mass tags, resolved on 4–12% Bis-Tris NuPAGE gels (Invitrogen), and transferred to nitrocellulose or PVDF membranes. The membranes were immunoblotted with antibodies against each protein of interest (see Supplementary Methods). After incubation with secondary antibodies (IRDye 800 goat anti-rabbit or Alexa Fluor 680 goat anti-mouse), proteins were visualized and quantified using an Odyssey infrared imaging system (LI-COR Biosciences). To quantify O-GlcNAc stoichiometries, the intensities of the PEG-shifted band (glycosylated protein fraction) and the unshifted band (non-glycosylated protein fraction) were measured using Odyssey imaging software (Version 2.1). The resulting values of the PEG-shifted bands were corrected for non-specific background by subtracting the background intensity from negative control reactions. For data and statistical analyses, mean values, standard error of the mean, and P-values (paired, two-tailed, Student’s T-tests, α-value = 0.05) were calculated using the program Excel.
Rexach J.E., Rogers C.J., Yu S.H., Tao J., Sun Y.E, & Hsieh-Wilson L.C. (2010). Quantitation of O-Glycosylation Stoichiometry and Dynamics using Resolvable Mass Tags. Nature chemical biology, 6(9), 645-651.
Publication 2010
Adult Animals Animals, Laboratory Antibodies Bistris Brain Cells Cortex, Cerebral Cultured Cells Cytosol Detergents Embryo Gels Glycosylated Proteins Goat HEK293 Cells Hypothalamus Institutional Animal Care and Use Committees IRDye800 Liver MECP2 protein, human Mus N-acetylglucosaminono-1,5-lactone O-(phenylcarbamoyl)oxime Neurons Nitrocellulose Pellets, Drug polyvinylidene fluoride Protein Glycosylation Proteins Rabbits Rats, Sprague-Dawley Seahorses Sf9 Cells Student Synapsins Tissue, Membrane
5
Quantifying Protein Glycosylation Dynamics
Cited 9 times
The following samples were used to detect glycosylation of the indicated proteins in Fig. 1c: 293T whole cell lysate (Sp1), rat hypothalamus crude nuclear pellet (MeCP2), rat brain detergent-soluble fraction (synapsin IIa, Nup62), whole cell lysate from cultured embryonic neurons (CREB), cytosolic fraction from PUGNAc-treated cultured embryonic neurons (OGA), and p75-OGT purified from Sf9 cells. For Fig. 3a, the liver, hippocampus or cerebral cortex was harvested from adult Sprague Dawley rats, and crude nuclear pellets were processed. Animal protocols were approved by the Institutional Animal Care and Use Committee at Caltech, and the procedures were performed in accordance with the Public Health Service Policy on Humane Care and Use of Laboratory Animals. Cell lysates were prepared as described in the Supplementary Methods. Each sample and its corresponding negative control (lacking ketogalactose probe 1 incorporation) was subjected to chemoenzymatic labeling with PEG mass tags, resolved on 4–12% Bis-Tris NuPAGE gels (Invitrogen), and transferred to nitrocellulose or PVDF membranes. The membranes were immunoblotted with antibodies against each protein of interest (see Supplementary Methods). After incubation with secondary antibodies (IRDye 800 goat anti-rabbit or Alexa Fluor 680 goat anti-mouse), proteins were visualized and quantified using an Odyssey infrared imaging system (LI-COR Biosciences). To quantify O-GlcNAc stoichiometries, the intensities of the PEG-shifted band (glycosylated protein fraction) and the unshifted band (non-glycosylated protein fraction) were measured using Odyssey imaging software (Version 2.1). The resulting values of the PEG-shifted bands were corrected for non-specific background by subtracting the background intensity from negative control reactions. For data and statistical analyses, mean values, standard error of the mean, and P-values (paired, two-tailed, Student’s T-tests, α-value = 0.05) were calculated using the program Excel.
Rexach J.E., Rogers C.J., Yu S.H., Tao J., Sun Y.E, & Hsieh-Wilson L.C. (2010). Quantitation of O-Glycosylation Stoichiometry and Dynamics using Resolvable Mass Tags. Nature chemical biology, 6(9), 645-651.
Publication 2010
Adult Animals Animals, Laboratory Antibodies Bistris Brain Cells Cortex, Cerebral Cultured Cells Cytosol Detergents Embryo Gels Glycosylated Proteins Goat HEK293 Cells Hypothalamus Institutional Animal Care and Use Committees IRDye800 Liver MECP2 protein, human Mus N-acetylglucosaminono-1,5-lactone O-(phenylcarbamoyl)oxime Neurons Nitrocellulose Pellets, Drug polyvinylidene fluoride Protein Glycosylation Proteins Rabbits Rats, Sprague-Dawley Seahorses Sf9 Cells Student Synapsins Tissue, Membrane

Most recents protocols related to «Glycosylated Proteins»

1
Optimized RBD Design via DMS Data

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Publication 2023
ACE2 protein, human Amino Acids Amino Acid Sequence Antibodies Base Sequence Binding Sites Cardiac Arrest Cysteine Epitopes Glycosylated Proteins Mutation Polysaccharides Saccharomyces cerevisiae
2
Molecular Docking of Curcumin and SARS-CoV-2
The molecular docking was conducted on the Lenovo ThinkPad T440p using the PyRx-Virtual Screening Tool. The structure of curcumin (sdf file format) was downloaded from the official website of the National Center for Biotechnology Information PubChem (https://pubchem.ncbi.nlm.nih.gov/, accessed on 3 January 2023). The energy minimization (optimization) was performed by a Universal Force Field (UFF). The 3D structure of curcumin–NLC constituents GMS, oleic acid, poloxamer 188, and tween 80 were obtained from drawing these chemical structures through ChemSchetch software and transferred from 2D to 3D and saved (as.mol file format). Then, they were converted to PDB format using the Discovery Studio visualizer 2019.
These structures were prepared for docking through PyRx software. The three host cell receptor structures of ACE2 (PDB ID: 7KMB, 7KNB, and 7KNH) were obtained from the RCSB PDB site (https://www.rcsb.org/, accessed on 3 January 2023). The receptor structures, with the aid of the Discovery Studio Visualizer 2021, were optimized, purified, and prepared for molecular docking. Autodock vina 1.1.2 in PyRx 0.8 was used to perform the molecular docking studies. A large number of glycosylated S proteins cover the surface of SARS-CoV-2 and bind to the host cell receptor angiotensin-converting enzyme 2 (ACE2), mediating a viral cell entry. Once the virus enters the cell, the viral RNA is released. Polyproteins are translated from the RNA genome, and the replication and transcription of the viral RNA genome occurs via the protein cleavage and assembly of the replicase–transcriptase complex. Viral RNA is replicated, and structural proteins are synthesized, assembled, and packaged in the host cell, after which viral particles are released [92 (link)]. For molecular docking, both ligands (PDBQT files), as well as the targets, were selected. The active binding receptors were examined and lightened using the BIOVIA Discovery Studio Visualizer (version-19.1.0.18287).
Albasri O.W., Kumar P.V, & Rajagopal M.S. (2023). Development of Computational In Silico Model for Nano Lipid Carrier Formulation of Curcumin. Molecules, 28(4), 1833.
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Publication 2023
Angiotensin Converting Enzyme 2 Cells Curcumin DNA Replication Genome Glycosylated Proteins Ligands Oleic Acid Poloxamer 188 Polyproteins Proteins Proteolysis RNA, Viral SARS-CoV-2 Transcriptase Transcription, Genetic Tween 80 Viral Genome Viral Transcription Virion Virus Virus Internalization
3
Formulation and Characterization of HIPEs
HIPEs were produced using a food processor (TM31 Thermomix, Vorwerk, Wuppertal, Germany), using 1% of PPI–MD conjugate (glycosylated protein) or PPI (control) for stabilizing the different formulations. First, PPI–MD conjugate or PPI was dispersed in 100 g of water and sheared for 30 s at 300 rpm (speed 3). Next, 400 g of oil was progressively introduced into the processor via gravitation from a separatory funnel and mixed at speed 3 for 3 min. The speed was then increased to 1100 rpm (speed 4) and kept until all oil was added. Finally, the emulsion was sheared for 30 s at 3100 rpm (speed 6). Six HIPEs were obtained following this procedure: 15A, 15B, 15P, 30A, 30B, and 30P. All the samples were prepared at least three times.
Morell P., López-García A., Hernando I, & Quiles A. (2023). Improving Pea Protein Emulsifying Capacity by Glycosylation to Prepare High-Internal-Phase Emulsions. Foods, 12(4), 870.
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Publication 2023
Emulsions Food Glycosylated Proteins Gravitation
4
Glycosylation of Duck Liver Protein
The glycosylated reaction of duck liver protein was prepared according to the method of Chen et al. [21 (link)] with some modifications. Duck liver protein was dissolved and sugars (glucose, fructose, and xylose) reduced in a beaker at the mass ratio of 1:4. The concentrations of duck liver protein and reducing sugar in the system were 1% and 4%, respectively. Then, it was magnetically stirred for 1 h and adjusted the pH of solution to 9.0 after being fully dissolved. Then, the mixture was put between duck liver protein and reducing sugar into the HH-8 digital constant temperature water bath (Lichen Bangxi Instrument Technology Co., Ltd., Shanghai, China) for 4 h at 363.15 K. It is worth noting that it should be sealed with plastic wrap to prevent the evaporation of the solution during the reaction. Finally, the glycosylated product of duck liver protein could be obtained when the thermal reaction was ended and cold. It should be noted that the reaction conditions were the optimal conditions for the glycosylated reaction of duck liver protein based on our previous research. The glycosylated product is sealed and stored at 277.15 K after lyophilized. In this study, the duck liver protein glycosylated products by glucose, fructose, and xylose were named as DLP-G, DLP-F, and DLP-X, respectively.
Zhan F., Luo J., Sun Y., Hu Y., Fan X, & Pan D. (2023). Antioxidant Activity and Cell Protection of Glycosylated Products in Different Reducing Sugar Duck Liver Protein Systems. Foods, 12(3), 540.
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Publication 2023
Adjustment Disorders Bath Cold Temperature Ducks Fingers Fructose Glucose Glycosylated Proteins Lichens Liver Proteins Sugars Xylose
5
Cytotoxicity Assessment of Duck Liver Glycosylated Products
The determination method of cytotoxicity tests was based on Hsu et al. [24 (link)] with minor modifications. The HepG2 cells were digested by 0.25% trypsin for 2 min when they reached 90% or more fusion. They were subculture at 1:2. We selected the logarithmic growth phase cells for experiment. The 100 mL of HepG2 cell suspension was at a density of 2 × 105 cells/mL per well into a 96 well culture plate and incubated at inoculated 310.15 K for 12 h. The original culture medium was discarded after the cells adhered and stabilized, and then 100 mL of cell culture medium containing duck liver protein glycosylated products was added. At this time, the final glycosylated products concentrations were 0.5, 1.0, 2.0, 2.5, 5.0, and 10.0 g/L. The cell viability was determined by method of CCK-8 after culture for 4 h. Each experimental group was set up with 5 replicates.
Zhan F., Luo J., Sun Y., Hu Y., Fan X, & Pan D. (2023). Antioxidant Activity and Cell Protection of Glycosylated Products in Different Reducing Sugar Duck Liver Protein Systems. Foods, 12(3), 540.
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Publication 2023
Cells Cell Survival Culture Media Cytotoxin Ducks Glycosylated Proteins Hepatocyte Hep G2 Cells Sincalide Trypsin

Top products related to «Glycosylated Proteins»

1
Pngase f by New England Biolabs
Sourced in United States, United Kingdom, Germany, Japan, Canada, France, China, Morocco, Italy
PNGase F is an enzyme that cleaves the bond between the asparagine residue and the N-acetylglucosamine residue in N-linked glycoproteins. It is commonly used in the analysis and characterization of glycoproteins.
2
Pro q emerald 300 glycoprot probes kombo by Thermo Fisher Scientific
Sourced in Italy
The PRO-Q EMERALD 300 GLYCOPROT PROBES KOMBO is a fluorescent stain used for the detection of glycoproteins in polyacrylamide gels. The probes specifically bind to glycoproteins, allowing their visualization and analysis.
3
Pro q emerald 300 glycoprotein gel and blot stain kit by Thermo Fisher Scientific
Sourced in United States
The Pro‐Q® Emerald 300 Glycoprotein Gel and Blot Stain Kit is a fluorescent stain used to detect glycoproteins in polyacrylamide gels and on Western blots. The kit contains all the necessary reagents for staining and visualization of glycoproteins.
4
Concanavalin a sepharose by Merck Group
Concanavalin A Sepharose is a chromatography resin composed of the lectin concanavalin A immobilized on Sepharose beads. It is used for the purification and isolation of glycoproteins and glycoconjugates from complex biological samples.
5
35s methionine by PerkinElmer
Sourced in United States
[35S]-methionine is a radioactive isotope of the amino acid methionine. It is commonly used as a labeling agent in various biochemical and molecular biology applications.
6
Biotin hydrazide by GE Healthcare
Biotin-hydrazide is a chemical compound used in various laboratory applications. It contains a biotin moiety and a hydrazide group, which allows it to react with carbonyl-containing compounds. Biotin-hydrazide is commonly used for the detection, purification, and immobilization of proteins, carbohydrates, and other biomolecules.
7
Sodium periodate by Merck Group
Sourced in United States, Germany, United Kingdom, China, Canada
Sodium periodate is a chemical compound with the formula NaIO4. It is a white, crystalline solid that is soluble in water. Sodium periodate serves as an oxidizing agent in various applications.
8
Schiff s reagent by Merck Group
Sourced in United States, Germany, United Kingdom, France
Schiff's reagent is a laboratory chemical used in various analytical and diagnostic applications. It is a solution composed of fuchsin, sulfurous acid, and hydrochloric acid. The primary function of Schiff's reagent is to detect the presence of aldehydes, which can be used to identify certain organic compounds and biomolecules.
9
Neuraminidase by New England Biolabs
Sourced in United States
Neuraminidase is an enzyme that catalyzes the removal of terminal sialic acid residues from glycoconjugates. It is commonly used in molecular biology and biochemistry applications.
10
Anti ha magnetic beads by Thermo Fisher Scientific
Sourced in United States, Germany
Anti-HA magnetic beads are a laboratory tool used for the isolation and purification of proteins tagged with the hemagglutinin (HA) epitope. These beads are coated with an antibody that specifically binds to the HA tag, allowing for the efficient capture and separation of HA-tagged proteins from complex biological samples.

More about "Glycosylated Proteins"

Glycosylated proteins, also known as glycoproteins, are a class of biomolecules where carbohydrate (sugar) groups are covalently attached to the protein backbone.
These complex structures play crucial roles in many biological processes, such as cell signaling, immune function, and protein folding.
Optimizing experimental protocols for working with glycosylated proteins is essential for advancing research in this field.
PubCompare.ai is an AI-driven platform that empowers researchers to locate, compare, and identify the most effective and reproducible methods from literature, preprints, and patents.
This tool simplifies the research process and enhances experimental results for studies involving glycosylated proteins.
Some key techniques and tools used in glycoprotein research include: - PNGase F: An enzyme that removes N-linked glycans from glycoproteins. - PRO-Q EMERALD 300 GLYCOPROT PROBES KOMBO: A fluorescent stain used to detect glycoproteins in polyacrylamide gels and Western blots. - Pro‐Q® Emerald 300 Glycoprotein Gel and Blot Stain Kit: Another glycoprotein staining solution. - Concanavalin A sepharose: A lectin-based affinity chromatography resin used to purify glycoproteins. - [35S]-methionine: A radioactive amino acid used to metabolically label and detect glycoproteins. - Biotin-hydrazide: A reagent that can be used to label and enrich for glycoproteins. - Sodium periodate: A chemical used to oxidize carbohydrates, which can be used to study glycan structures. - Schiff's reagent: A stain that can be used to detect carbohydrates, including those found in glycoproteins. - Neuraminidase: An enzyme that removes sialic acid residues from glycoproteins. - Anti-HA magnetic beads: A tool used to immunoprecipitate and purify HA-tagged glycoproteins.
By leveraging the insights and tools available for glycoprotein research, scientists can optimize their experimental protocols and enhance the quality and reproducibility of their studies.
PubCompare.ai provides a valuable resource to help researchers navigate this field and identify the most effective methods.