To increase the probability of successful crystallization and structure determination, several soluble, cleaved trimers of the SOSIP.664 gp140 design were expressed and purified as described elsewhere (17 (link), 18 (link), 23 (link), 64 (link)). Crystallization candidates based on env genes from HIV-1 isolates KNH1144, ADA, 1182.6.1D2, BG505 and 208.9.C10 were selected from an initial panel of 20, based on their expression and trimer-formation properties, as assessed by SDS-PAGE and BN-PAGE. Notably, for isolates that did not naturally contain residue N332, a point substitution was made to introduce it, thereby creating the 2G12 epitope to facilitate affinity purification. The Env constructs were co-transfected with the furin protease in HEK 293S GnTI−/− cells, which lack N-acetylglucosaminyltransferase I and, therefore, produce glycoproteins bearing only high-mannose (Man5-9) glycans. Secreted SOSIP.664 Env proteins were harvested from supernatants and affinity purified using a 2G12 MAb affinity column. Following a high salt elution process, trimers were purified to size homogeneity using a Superose 6 size exclusion chromatography (SEC) matrix (GE Healthcare).
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Mannose
Mannose
Mannose is a monosaccharide sugar that is an important component of glycoproteins and glycolipids in the body.
It plays a key role in various biological processes, such as cell signaling, immune function, and protein folding.
Mannose can be obtained from dietary sources, such as fruits and vegetables, or synthesized endogenously.
Researchers are actively investigating the potential therapeutic applications of mannose, including its use in the treatment of infections, cancer, and metabolic disorders.
PubCompare.ai can help streamline your mannose research by providing access to the latest protocols and literature, enabling you to identify the most effective and reproducible approaches.
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It plays a key role in various biological processes, such as cell signaling, immune function, and protein folding.
Mannose can be obtained from dietary sources, such as fruits and vegetables, or synthesized endogenously.
Researchers are actively investigating the potential therapeutic applications of mannose, including its use in the treatment of infections, cancer, and metabolic disorders.
PubCompare.ai can help streamline your mannose research by providing access to the latest protocols and literature, enabling you to identify the most effective and reproducible approaches.
Experienec the power of AI-driven research optimization today.
Most cited protocols related to «Mannose»
alpha-1,3-mannosyl-glycoprotein beta-1,2-N-acetylglucosaminyltransferase I
Cells
Chromatography, Affinity
Crystallization
Epitopes
FURIN protein, human
Gel Chromatography
Gene Products, env
Glycoproteins
GP 140
HIV-1
Mannose
Peptide Hydrolases
Polysaccharides
Salts
SDS-PAGE
Genesets of interest were identified by the consortium and separated in five main groups, as detailed in Supplementary Table 9 and below:
ESTIMATE algorithm: method that uses gene expression signatures to infer the fraction of stromal and immune cells in tumor samples30 (link);
Curated signatures: upper and lower normal colon crypt compartments51 , epithelial and mesenchymal markers7 (link), WNT52 and MYC downstream target53 , epithelial-mesenchymal transition core genes and TGFβ pathway54 , intestinal stem cells55 , matrix remodeling (REACTOME) and wound-response (GO BP);
Canonical genesets: MAPK and PI3K (GO BP), SRC, JAK-STAT, caspases (BIOCARTA), proteosome (KEGG), Notch, cell cycle, translation and ribosome, integrin beta3, VEGF/VEGFR interactions (REACTOME);
Immune activation: immune response (GO BP), PD1 activation (REACTOME), infiltration with T cytotoxic cells (CD8)56 and T helper cells (TH1) in cancer samples57 ,58 , infiltration with Natural Killer (NK) cells59 and follicular helper T (TFH) cells60 in cancer samples, activation of T helper 17 (TH17) cells61 , regulatory T cells (Treg)62 or myeloid-derived suppressor cells (MDSC)63 ;
Metabolic activation: sugar, amino acid, nucleotide, glucose, pentose, fructose, mannose, starch, sucrose, galactose, glutathione, nitrogen, tyrosine, glycerophospholipid, fatty acid, arachnoid acid, linoleic acid (KEGG), glutamine (GO BP), lysophospholipid (PID).
Acids
Activation, Metabolic
Amino Acids
Arachnoid Maters
Carbohydrates
Caspase
Cell Cycle
Cells
CFC1 protein, human
Colon
Cytotoxic T-Lymphocytes
Fatty Acids
FLT1 protein, human
Fructose
Galactose
Genes
Glucose
Glutamine
Glutathione
Glycerophospholipids
Helper-Inducer T-Lymphocyte
Integrin beta3
Intestines
Linoleic Acid
Lysophospholipids
Malignant Neoplasms
Mannose
Mesenchyma
Multicatalytic Endopeptidase Complex
Myeloid-Derived Suppressor Cells
Neoplasms
Nitrogen
Nucleotides
Pentoses
Phosphatidylinositol 3-Kinases
Regulatory T-Lymphocytes
Response, Immune
Ribosomes
Starch
Stem, Plant
Sucrose
Transforming Growth Factor beta
Transition, Epithelial-Mesenchymal
Tyrosine
Vascular Endothelial Growth Factors
Wounds
11-cis-Retinal
Biological Assay
Bos taurus
Cells
Chromatography, Affinity
Dithionite
Fatty Acids
Fluorescence
Glycolipids
Homo sapiens
Lipids
Mannose
Membrane Proteins
Monoclonal Antibodies
Phospholipids
proteoliposomes
Rhodopsin
Rod Cell Outer Segment
Rod Opsins
SDS-PAGE
Serum Albumin
Triton X-100
Env proteins were purified from the supernatants by affinity chromatography using either a 2G12 column or a Galanthus nivalis (GN)-lectin column [25] (link), [27] (link), [46] (link), [64] (link). Briefly, transfection supernatants were vacuum filtered through 0.2-µm filters and then passed (0.5–1 ml/min flow rate) over the column. The 2G12 column was made from CNBr-activated Sepharose 4B beads (GE Healthcare) coupled to the bNAb 2G12 (Polymun Sciences, Klosterneuburg, Austria). Purification using this column was performed as follows: the beads were washed with 2 column volumes of buffer (0.5 M NaCl, 20 mM Tris, pH 8.0) before eluting bound Env proteins using 1 column volume of 3 M MgCl2. The eluted proteins were immediately buffer exchanged into 75 mM NaCl, 10 mM Tris, pH 8.0, using Snakeskin dialysis tubing (10K WCMO) (Thermo Scientific). The buffer-exchanged proteins were further concentrated using Vivaspin columns with a 30-kDa cut off (GE Healthcare). For GN-lectin affinity purification, the wash buffer was Dulbecco's phosphate buffer saline (DPBS) supplemented with 0.5 M NaCl was used, and elution was carried out using DPBS supplemented with 1 M methyl mannopyranoside.
In both cases, the affinity-purified Env proteins were further purified to size homogeneity using size exclusion chromatography (SEC) on a Superdex 200 26/60 column (GE Healthcare). A Superose 6 column was sometimes used for analytical or preparative purposes. The trimer fractions and, occasionally also the SOSIP gp140 monomer fractions, were collected and pooled. Protein concentrations were determined using either a bicinchonic acid-based assay (BCA assay; Thermo Scientific, Rockford, IL) or UV280 absorbance using theoretical extinction coefficients [66] .
In both cases, the affinity-purified Env proteins were further purified to size homogeneity using size exclusion chromatography (SEC) on a Superdex 200 26/60 column (GE Healthcare). A Superose 6 column was sometimes used for analytical or preparative purposes. The trimer fractions and, occasionally also the SOSIP gp140 monomer fractions, were collected and pooled. Protein concentrations were determined using either a bicinchonic acid-based assay (BCA assay; Thermo Scientific, Rockford, IL) or UV280 absorbance using theoretical extinction coefficients [66] .
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Acids
Biological Assay
Broadly Neutralizing Antibodies
Buffers
Chromatography, Affinity
Cyanogen Bromide
Dialysis
Extinction, Psychological
Gel Chromatography
Gene Products, env
GP 140
Magnesium Chloride
Mannose
Phosphates
Proteins
Saline Solution
Sepharose 4B
snowdrop lectin
Sodium Chloride
Transfection
Tromethamine
Vacuum
Antibody Affinity
Cells
Chromatography, Affinity
Cloning Vectors
Gel Chromatography
Genes, env
GP 140
Mannose
Proteins
SDS-PAGE
Sepharose
snowdrop lectin
Sodium Chloride
Transfection
Most recents protocols related to «Mannose»
Example 3
The genes for Candida antartica lipases A and B, human transferrin, and the human CH2 domain from IgG were integrated into the SuperM5 genome using standard transformation methods. In all cases significant amounts of protein were produced and secreted into the medium. Transformed strains and media-containing protein were tested for glycan analysis using previously published methods. In all cases, the glycan profiles for the test proteins and for the strain glycoproteins demonstrated a mannose-5 glycan structure with no other higher mannose structures detected by the methods used.
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Candidiasis, Genital
Genome
Glycoproteins
Homo sapiens
Lipase
Mannose
Polysaccharides
Proteins
Strains
Transferrin
Example 2
SuperM5 was stored in different conditions at −80° C., −4° C., 20° C. and at room temperature. Strains were stored as frozen glycerol stocks and as stab cultures. Different cultures were stored and thawed for different experiments and for shipping to collaborators for testing. In all cases the strains recovered, plated and cultured similar to the parent Pichia pastoris GS115 strain and grew in both complex and defined media similar to the parent strains. The SuperM5 strains transformed similarly as the parent strain and proteins were expressed with the mannose-5 glycosylation as the predominate glycoform, or the only glycoform. Strains have been repeatedly stored and regrown to establish robustness of the SuperM5 strains.
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Freezing
Glycerin
Komagataella pastoris
Mannose
Parent
Protein Glycosylation
Proteins
Strains
Molecular weight distributions of lyophilized crude EPS were determined by size exclusion chromatography. In brief, crude EPS powder was suspended in 0.1 M NaNO3 (0.5 mg/mL) and then filtered through a 0.45 μm pore diameter polyvinylidene fluoride membrane (Millipore Corporation, USA). The average molecular weight (MW) was determined by high-performance molecular exclusion chromatography (HPLC-SEC, Agilent 1,100 Series System, Hewlett-Packard, Germany) associated with a refractive index (IR) detector (Ibarburu et al., 2015 (link)). 50 μL of the samples were injected and eluted at a flow rate of 0.95 mL/min (pressure: 120:130 psi) at room temperature using 0.1 M NaNO3 as mobile phase. Dextrans (0.5 mg/mL) with a molecular weight between 103 and 2.106 Da (Sigma-Aldrich, USA) were used as standards.
Once the molecular weight distributions were determined, low and high molecular weight fractions that composed the crude EPS obtained at 20°C were separated. For this purpose, EPS solutions (0.2% w/v) were centrifuged through a Vivaspin™ ultrafiltration spin column 100 KDa MWCO, (Sartorious, Goettingen, Germany) for 20 min at 6000 g, eluting only the low MW fraction. Subsequently, high MW fraction retained in the column was eluted using hot distilled water. The eluted fractions were passed through a Vivaspin column (cut-off 30KDa) in order to separate the middle and low MW fraction of EPS.
Monosaccharide composition of crude EPS and their fractions were determined by gas chromatography as previously described (Notararigo et al., 2013 (link)). Briefly, 1–2 mg of EPS were hydrolyzed in 1 mL of 3 M trifluoroacetic acid (1 h at 120°C). The monosaccharides obtained were converted into alditol acetates by reduction with NaBH4 and subsequent acetylation. The samples were analyzed by gas chromatography in an Agilent 7890A coupled to a 5975C mass detector, using an HP5-MS column with helium as carrier gas at a flow rate of 1 mL/min. For each run, 1 μL of sample was injected (with a Split 1:50) and the following temperature program was performed: the oven was heat to 175°C for 1 min; the temperature was increased to 215°C at a rate of 2.5°C/min and then increased to 225°C at 10°C/min, keeping it constant at this temperature for 1.5 min. Monosaccharides were identified by comparison of retention times with standards (arabinose, xylose, rhamnose, galactose, glucose, mannose, glucosamine and galactosamine) analyzed under the same conditions. Calibration curves were also processed for monosaccharide quantification. Myo-inositol was added to each sample as internal standard.
Once the molecular weight distributions were determined, low and high molecular weight fractions that composed the crude EPS obtained at 20°C were separated. For this purpose, EPS solutions (0.2% w/v) were centrifuged through a Vivaspin™ ultrafiltration spin column 100 KDa MWCO, (Sartorious, Goettingen, Germany) for 20 min at 6000 g, eluting only the low MW fraction. Subsequently, high MW fraction retained in the column was eluted using hot distilled water. The eluted fractions were passed through a Vivaspin column (cut-off 30KDa) in order to separate the middle and low MW fraction of EPS.
Monosaccharide composition of crude EPS and their fractions were determined by gas chromatography as previously described (Notararigo et al., 2013 (link)). Briefly, 1–2 mg of EPS were hydrolyzed in 1 mL of 3 M trifluoroacetic acid (1 h at 120°C). The monosaccharides obtained were converted into alditol acetates by reduction with NaBH4 and subsequent acetylation. The samples were analyzed by gas chromatography in an Agilent 7890A coupled to a 5975C mass detector, using an HP5-MS column with helium as carrier gas at a flow rate of 1 mL/min. For each run, 1 μL of sample was injected (with a Split 1:50) and the following temperature program was performed: the oven was heat to 175°C for 1 min; the temperature was increased to 215°C at a rate of 2.5°C/min and then increased to 225°C at 10°C/min, keeping it constant at this temperature for 1.5 min. Monosaccharides were identified by comparison of retention times with standards (arabinose, xylose, rhamnose, galactose, glucose, mannose, glucosamine and galactosamine) analyzed under the same conditions. Calibration curves were also processed for monosaccharide quantification. Myo-inositol was added to each sample as internal standard.
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Acetates
Acetylation
Arabinose
Dextrans
Division Phase, Cell
Galactosamine
Galactose
Gas Chromatography
Gel Chromatography
Glucosamine
Glucose
Helium
High-Performance Liquid Chromatographies
Inositol
Mannose
Monosaccharides
polyvinylidene fluoride
Powder
Pressure
Retention (Psychology)
Rhamnose
Sugar Alcohols
Tissue, Membrane
Trifluoroacetic Acid
Ultrafiltration
Xylose
According to the results of transcriptomic and proteomic analyses, the carbohydrate substance (mannose, galactose, cellobiose, and D-ribose) was added to the mono-culture of L. paraplantarum RX-8 at the concentrations of 2, 20, and 200 mM. Meanwhile, the amino acid substance (arginine, cysteine, glutamate, and glutamine) was added to the mono-culture of L. paraplantarum RX-8 at the concentrations of 0.5, 1.25, and 2.5 g/L. After being cultured for 24 h at 37°C, the supernatants of each sample were collected for the plantaricin production assay. The mono-culture of L. paraplantarum RX-8 was used as a control.
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Amino Acids
Arginine
Biological Assay
Carbohydrates
Cellobiose
Cysteine
Galactose
Gene Expression Profiling
Glutamates
Glutamine
Mannose
Ribose
The process of labeling experiments followed a previous protocol (Wang et al., 2020 (link)). For mannose labeling, ∼2 × 106 PIGS-KO, PIGS-HRD1-DKO, and PIGS-HRD1-CD55-TKO cells, and PIGS-HRD1-CD55-TKO cells stably expressing HA-CD55 were precultured in normal medium overnight, washed with wash medium (glucose-free DMEM buffered with 20 mM Hepes, pH 7.4), and incubated for 1 h at 37°C in 1 ml of reaction medium (wash medium supplemented with 10% dialyzed FBS (Gibco), 10 μg ml−1 tunicamycin (Wako), and 100 μg ml−1 glucose). [2-3H] Mannose (American Radiolabeled Chemicals) was added to 25 Ci ml−1 and the cells were incubated for 1 h at 37°C in 5% CO2. The cells were pelleted and washed with 1 ml of cold PBS. Radiolabeled GPIs were extracted with 1-butanol, separated by high-performance thin-layer chromatography (HPTLC; Merck), and visualized using an FLA 7000 analyzer (Fujifilm).
To detect GlcN-PI or GlcN-(acyl)-PI after inositol-labeling, PIGW-KO, PIGS-KO, PIGS-HRD1-DKO, PIGS-HRD1-ARV1-TKO, PIGS-HRD1-CD55-TKO, and HA-CD55 rescued cells were washed with inositol-free DMEM and then incubated in 1 ml of reaction medium B (inositol-free DMEM buffered with 20 mM Hepes, pH 7.4) supplemented with 10% dialyzed FBS in the presence of 10 μCi of myo-[2-3H] inositol (PerkinElmer) for 24 h. After metabolic labeling, the cells were washed twice with 1 ml of cold PBS and pelleted by centrifugation. Lipids and radiolabeled GPIs were extracted with 1-butanol partitioning, separated by HPTLC (Merck), and visualized using an FLA 7000 analyzer.
To detect GlcN-PI or GlcN-(acyl)-PI after inositol-labeling, PIGW-KO, PIGS-KO, PIGS-HRD1-DKO, PIGS-HRD1-ARV1-TKO, PIGS-HRD1-CD55-TKO, and HA-CD55 rescued cells were washed with inositol-free DMEM and then incubated in 1 ml of reaction medium B (inositol-free DMEM buffered with 20 mM Hepes, pH 7.4) supplemented with 10% dialyzed FBS in the presence of 10 μCi of myo-[2-3H] inositol (PerkinElmer) for 24 h. After metabolic labeling, the cells were washed twice with 1 ml of cold PBS and pelleted by centrifugation. Lipids and radiolabeled GPIs were extracted with 1-butanol partitioning, separated by HPTLC (Merck), and visualized using an FLA 7000 analyzer.
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Butyl Alcohol
Cells
Centrifugation
Cold Temperature
Glucose
HEPES
Inositol
Lipids
Mannose
Pigs
Thin Layer Chromatography
Tunicamycin
Top products related to «Mannose»
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D-mannose is a type of sugar that can be used as a component in laboratory equipment and processes. It serves as a basic chemical substance for various applications in research and development.
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Mannose is a type of sugar molecule that is commonly used in laboratory settings. It serves as a core structural component in various biological compounds and can be utilized in a variety of applications within the scientific research field.
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Galactose is a monosaccharide that serves as a core component in various laboratory analyses and experiments. It functions as a fundamental building block for complex carbohydrates and is utilized in the study of metabolic processes and cellular structures.
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D-glucose is a type of monosaccharide, a simple sugar that serves as the primary source of energy for many organisms. It is a colorless, crystalline solid that is soluble in water and other polar solvents. D-glucose is a naturally occurring compound and is a key component of various biological processes.
Sourced in United States, Germany, China, Switzerland, Sao Tome and Principe, Spain, United Kingdom, Ireland, Sweden
Xylose is a monosaccharide that can be used in laboratory equipment and procedures. It is a key component in various biochemical and analytical applications.
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Arabinose is a monosaccharide that is commonly used as a component in various laboratory equipment and supplies. It functions as a carbohydrate source and can be utilized in various biochemical and microbiological applications.
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D-galactose is a monosaccharide carbohydrate. It is a constituent of many natural polysaccharides, including lactose, cerebrosides, and gangliosides. D-galactose can be used as a laboratory reagent.
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Rhamnose is a monosaccharide that serves as a core component in various glycoconjugates. It is a sugar alcohol commonly used in biochemical and microbiological applications as a carbon source and for the cultivation of certain bacteria and fungi.
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D-xylose is a monosaccharide sugar that can be used in various laboratory applications. It is a pentose sugar that is naturally found in plant materials. D-xylose has a wide range of potential uses in research and analysis, but a detailed description of its core function is not available while maintaining an unbiased and factual approach.
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Fucose is a monosaccharide commonly found in glycoproteins and glycolipids. It serves as a core component in various biological processes.
More about "Mannose"
Mannose, also known as D-mannose or hexose, is a versatile monosaccharide sugar that plays a crucial role in various biological processes within the human body.
As a component of glycoproteins and glycolipids, mannose is essential for cell signaling, immune function, and protein folding.
This sugar can be obtained from dietary sources, such as fruits and vegetables, or synthesized endogenously through metabolic pathways.
Beyond its fundamental biological functions, researchers are actively investigating the potential therapeutic applications of mannose.
Its use in the treatment of infections, cancer, and metabolic disorders is an area of growing interest.
Closely related monosaccharides, such as galactose, glucose, xylose, arabinose, and rhamnose, also exhibit unique properties and roles in the body, making them subject to extensive scientific exploration.
To streamline your mannose research, PubCompare.ai offers an AI-driven platform that helps you locate the best protocols from literature, pre-prints, and patents.
By providing accurate comparisons, the tool enables you to identify the most reproducible and effective approaches, empowering you to uncover the insights you need with just a few clicks.
Experience the future of research optimization and unlock the full potential of your mannose studies today!
As a component of glycoproteins and glycolipids, mannose is essential for cell signaling, immune function, and protein folding.
This sugar can be obtained from dietary sources, such as fruits and vegetables, or synthesized endogenously through metabolic pathways.
Beyond its fundamental biological functions, researchers are actively investigating the potential therapeutic applications of mannose.
Its use in the treatment of infections, cancer, and metabolic disorders is an area of growing interest.
Closely related monosaccharides, such as galactose, glucose, xylose, arabinose, and rhamnose, also exhibit unique properties and roles in the body, making them subject to extensive scientific exploration.
To streamline your mannose research, PubCompare.ai offers an AI-driven platform that helps you locate the best protocols from literature, pre-prints, and patents.
By providing accurate comparisons, the tool enables you to identify the most reproducible and effective approaches, empowering you to uncover the insights you need with just a few clicks.
Experience the future of research optimization and unlock the full potential of your mannose studies today!