100 assays (manual) / 1000 assays (microplate) / 1020 assays (auto-analyser)
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|Content:||100 assays (manual) / 1000 assays (microplate) / 1020 assays (auto-analyser)|
Short term stability: 2-8oC,
Long term stability: See individual component labels
|Stability:||> 2 years under recommended storage conditions|
|Assay Format:||Spectrophotometer, Microplate, Auto-analyser|
|Linear Range:||0.5 to 100 µg of L-fucose per assay|
|Limit of Detection:||0.68 mg/L|
|Reaction Time (min):||~ 10 min|
|Application examples:||L-Fucose is present as the main component in fucoidan (a marine polysaccharide), foods, pharmaceuticals and other materials (e.g. biological samples, etc.).|
|Method recognition:||Novel method|
The L-Fucose test kit is a simple, rapid and reliable method, for the measurement and analysis of L-Fucose in plant extracts, biological samples and other materials. This kit can be used in the measurement of α-fucosidases that do not act on chromogenic substrates.
Note for Content: The number of manual tests per kit can be doubled if all volumes are halved. This can be readily accommodated using the MegaQuantTM Wave Spectrophotometer (D-MQWAVE).
See our full range of monosaccharide and disaccharide assay kits.
- Very cost effective
- All reagents stable for > 2 years after preparation
- Only enzymatic kit available
- Simple format
- Rapid reaction time (~ 10 min)
- Mega-Calc™ software tool is available from our website for hassle-free raw data processing
- Standard included
- Suitable for manual, microplate and auto-analyser formats
Production of perdeuterated fucose from glyco-engineered bacteria.
Gajdos, L., Forsyth, V. T., Blakeley, M. P., Haertlein, M., Imberty, A., Samain, E. & Devos, J. M. (2020). Glycobiology, In Press.
L-Fucose and L-fucose-containing polysaccharides, glycoproteins or glycolipids play an important role in a variety of biological processes. L-Fucose-containing glycoconjugates have been implicated in many diseases including cancer and rheumatoid arthritis. Interest in fucose and its derivatives is growing in cancer research, glyco-immunology, and the study of host–pathogen interactions. L-Fucose can be extracted from bacterial and algal polysaccharides or produced (bio)synthetically. While deuterated glucose and galactose are available, and are of high interest for metabolic studies and biophysical studies, deuterated fucose is not easily available. Here, we describe the production of perdeuterated L-fucose, using glyco-engineered Escherichia coli in a bioreactor with the use of a deuterium oxide-based growth medium and a deuterated carbon source. The final yield was 0.2 g L−1 of deuterated sugar, which was fully characterized by mass spectrometry and nuclear magnetic resonance spectroscopy. We anticipate that the perdeuterated fucose produced in this way will have numerous applications in structural biology where techniques such as NMR, solution neutron scattering and neutron crystallography are widely used. In the case of neutron macromolecular crystallography, the availability of perdeuterated fucose can be exploited in identifying the details of its interaction with protein receptors and notably the hydrogen bonding network around the carbohydrate binding site.Hide Abstract
Fucosidases from the human gut symbiont Ruminococcus gnavus.
Wu, H., Rebello, O., Crost, E. H., Owen, C. D., Walpole, S., Bennati-Granier, C., Ndeh, D., Monaco, S., Hicks, T., Colvile, A., A. Urbanowicz, P., Walsh, M. A., Angulo, J., Spencer, D. I. R. & Juge, N. (2020). Cellular and Molecular Life Sciences, 1-19.
The availability and repartition of fucosylated glycans within the gastrointestinal tract contributes to the adaptation of gut bacteria species to ecological niches. To access this source of nutrients, gut bacteria encode α-L-fucosidases (fucosidases) which catalyze the hydrolysis of terminal α-L-fucosidic linkages. We determined the substrate and linkage specificities of fucosidases from the human gut symbiont Ruminococcus gnavus. Sequence similarity network identified strain-specific fucosidases in R. gnavus ATCC 29149 and E1 strains that were further validated enzymatically against a range of defined oligosaccharides and glycoconjugates. Using a combination of glycan microarrays, mass spectrometry, isothermal titration calorimetry, crystallographic and saturation transfer difference NMR approaches, we identified a fucosidase with the capacity to recognize sialic acid-terminated fucosylated glycans (sialyl Lewis X/A epitopes) and hydrolyze α1-3/4 fucosyl linkages in these substrates without the need to remove sialic acid. Molecular dynamics simulation and docking showed that 3′-Sialyl Lewis X (sLeX) could be accommodated within the binding site of the enzyme. This specificity may contribute to the adaptation of R. gnavus strains to the infant and adult gut and has potential applications in diagnostic glycomic assays for diabetes and certain cancers.Hide Abstract
Engineering two species of yeast as cell factories for 2′-fucosyllactose.
Hollands, K., Baron, C. M., Gibson, K. J., Kelly, K. J., Krasley, E. A., Laffend, L. A., Lauchli, R. M., Maggio-Hall, L. A., Nelson, M. J., Prasad, J. C., Ren, Y., Rice, B. A., Rice, G. H. & Rothman, S. C. (2019). Metabolic Engineering, 52, 232-242.
Oligosaccharides present in human breast milk have been linked to beneficial effects on infant health. Inclusion of these human milk oligosaccharides (HMOs) in infant formula can recapitulate these health benefits. As a result, there is substantial commercial interest in a cost-effective source of HMOs as infant formula ingredients. Here we demonstrate that the yeast species Saccharomyces cerevisiae and Yarrowia lipolytica both can be engineered to produce 2′-fucosyllactose (2′FL), which is the most abundant oligosaccharide in human breast milk, at high titer and productivity. Both yeast species were modified to enable uptake of lactose and synthesis of GDP-fucose - the two precursors of 2′FL - by installing a lactose transporter and enzymes that convert GDP-mannose to GDP-fucose. Production of 2′FL was then enabled by expression of α-1,2-fucosyltransferases from various organisms. By screening candidate transporters from a variety of sources, we identified transporters capable of exporting 2′FL from yeast, which is a key consideration for any biocatalyst for 2′FL production. In particular, we identified CDT2 from Neurospora crassa as a promising target for further engineering to improve 2′FL efflux. Finally, we demonstrated production of 2′FL in fermenters at rates and titers that indicate the potential of engineered S. cerevisiae and Y. lipolytica strains for commercial 2′FL production.Hide Abstract
Development of a quantitative assay for 2´-fucosyllactose via one-pot reaction with α1, 2-fucosidase and L-fucose dehydrogenase.
Seydametova, E., Shin, J., Yu, S. H., Kim, C., Kim, H., Park, Y. J., Yang, J. K., Cho, S., Kim, H. R., Moon, S., Ban, C. & Kweon, D. H. (2019). Analytical Biochemistry, 582, 113358.
2′-Fucosyllactose (2′-FL) is the most abundant milk oligosaccharide in human breast milk and it has several benefits for infant health. The quantification of 2′-FL in breast milk or in samples from other sources generally requires lengthy analyses. These methods cannot be used to simultaneously detect 2′-FL in numerous samples, which would be more time-efficient. In this study, two genes, namely α1,2-fucosidase from Xanthomonas manihotis and L-fucose dehydrogenase from Pseudomonas sp. no. 1143, were identified, cloned and overexpressed in E. coli. The recombinant enzymes were produced as 6 × His-tagged proteins and were purified to homogeneity using Ni2+ affinity chromatography. The purified α1,2-fucosidase and l-fucose dehydrogenase are monomers with molecular masses of 63 kDa and 36 kDa, respectively. Both enzymes have sufficiently high activities in phosphate-buffered saline (pH 7.0) at 37°C, making it possible to develop a coupled enzyme reaction in a single buffer system for the quantitative determination of 2′-FL in a large number of samples simultaneously. This method can be used to quantify 2′-FL in infant formulas and in samples collected from different phases of the biotechnological production of this oligosaccharide. Furthermore, the method is applicable for the rapid screening of active variants during the development of microbial strains producing 2′-FL.Hide Abstract
Interactions of anthocyanins with pectin and pectin fragments in model solutions.
Larsen, L. R., Buerschaper, J., Schieber, A. & Weber, F. (2019). Journal of Agricultural and Food Chemistry, 67(33), 9344-9353.
Anthocyanins determine the color and potential health-promoting properties of red fruit juices, but the juices contain remarkably less anthocyanins than the fruits, which is partly caused by the interactions of anthocyanins with the residues of cell wall polysaccharides like pectin. In this study, pectin was modified by ultrasound and enzyme treatments to residues of polysaccharides and oligosaccharides widely differing in their molecular weight. Modifications decreased viscosity and degrees of acetylation and methylation and released smooth and hairy region fragments. Native and modified pectin induced different effects on the concentrations of individual anthocyanins after short-term and long-term incubation caused by both hydrophobic and hydrophilic interactions. Results indicate that both pectin and anthocyanin structure influence these interactions. Linear polymers generated by ultrasound formed insoluble anthocyanin complexes, whereas oligosaccharides produced by enzymes formed soluble complexes with protective properties. The structure of the anthocyanin aglycone apparently influenced interactions more than the sugar moiety.Hide Abstract
Assaying fucosidase activity.
Megson, Z. A., Messner, P. & Schäffer, C. (2019). Bacterial Polysaccharides, Humana Press, New York, NY, 269-278.
The characterization of a recombinant glycosidase can be done with commercially available substrates, which enable testing of enzyme functionality and determination of linkage specificity. Colorimetric assays with p-nitrophenyl substrates provide a relatively simple and fast way of screening conditions which could affect enzyme activity (buffer, pH, ion dependence, temperature). These substrates are useful for the determination of activity optima and the characterization of basic activity parameters. However, testing for linkage specificity should be performed on more complex sugars presenting a range of different glycosidic bonds and might need more sophisticated methods of analysis. This protocol provides comprehensive instructions on how to perform an initial characterization of your glycosidase using a recombinant α-l-fucosidase as an example.Hide Abstract
Cross-linking of diluted alkali-soluble pectin from apple (Malus domestica fruit) in different acid-base conditions.
Gawkowska, D., Cieśla, J., Zdunek, A. & Cybulska, J. (2019). Food Hydrocolloids, 92, 285-292.
A diluted alkali-soluble pectin (DASP) fraction, extracted using sodium carbonate, is characterized by a low degree of methylesterification and has the ability to self-organize on mica. The aim of this study was to characterize the cross-linking process of this fraction, extracted from apples, over a wide pH range (3-11) and without the addition of salt. An FT-IR study showed an increase in the intensity of bands connected with νas and νs (COO−) and a decrease in the intensity of the band associated with ν (C=O) in the carboxyl group with increasing pH, which indicated the dissociation of the carboxyl groups of galacturonic acid units. An increase in the surface electrical charge of particles in the pH range of 3-7 confirmed this. The value of the average apparent dissociation constant (∼4.60) indicated the acidic character of the DASP fraction. An AFM study showed the morphological changes of the DASP fraction with increasing pH, which allowed for the evaluation of the cross-linking process. This fraction formed a network on mica at pH 4 and 9, while the aggregates were noted mainly at pH 11. For totally ionized carboxyl groups (pH 7), the pectin chains were separated from each other due to the electrostatic repulsion between them.Hide Abstract
Ustyuzhanina, N. E., Bilan, M. I., Dmitrenok, A. S., Borodina, E. Y., Nifantiev, N. E. & Usov, A. I. (2018). Carbohydrate Research, 456, 5-9.
A highly regular fucan sulfate SHFS was isolated from the sea cucumber Stichopus horrens by extraction of the body walls in the presence of papain followed by ion-exchange and gel permeation chromatography. SHFS had MW of about 140 kDa and contained fucose and sulfate in the molar ratio of about 1:1. Chemical and NMR spectroscopic methods were applied for the structural characterization of the polysaccharide. SHFS was shown to have linear molecules built up of 3-linked α-L-fucopyranose 2-sulfate residues. Anticoagulant properties of SHFS were assessed in vitro in comparison with the LMW heparin (enoxaparin) and totally sulfated 3-linked α-L-fucan. SHFS was found to have the lowest activity, and hence, both sulfate groups at O-2 and O-4 of fucosyl units seem to be important for anticoagulant effect of sulfated homo-(1 → 3)-α-L-fucans.Hide Abstract
Martínez–Hernández, G. B., Castillejo, N., Carrión–Monteagudo, M. D. M., Artés, F. & Artés-Hernández, F. (2017). Food Science and Technology International, 1082013217740000.
The main nutritional/bioactive compounds (protein; aminoacids, AA; fucose; minerals; vitamins B12 and C; and total phenolic content, TPC) of nine commercial algae powders, used as food supplements, were studied. Undaria pinnatifida showed the highest protein/aminoacid contents (51.6/54.4 g 100 g-1). Among brown macroalgae, Himanthalia elongate showed the highest fucose content (26.3 g kg-1) followed by Laminaria ochroleuca (22.5 g kg-1). Mineral contents of 15-24% were observed in the algae, being particularly excellent sources of iodine (69.0-472.0 mg kg-1). Porphyra spp. and Palmaria palmata showed the highest vitamin B12 contents (667-674 µg kg-1). Vitamin C ranged among 490.4-711.8 mg kg-1. H. elongate showed the highest total phenolic content (14.0 g kg-1). In conclusion, the studied algae are excellent sources of protein, AA, minerals, vitamin C and some of them presented particularly high vitamin B12 and fucose contents, which may have a potential use as food supplements.Hide Abstract
Manchil, P. R. D., Joy, E. T., Kiran, M. S., Sherubin, J. E., Khan, M. F. & Aravind, B. S. (2016). Journal of Pharmacy & Bioallied Sciences, 8(Suppl 1), S147-S150.
Background: oral cancer is a result of disordered cellular behavior initiated by various stimuli which is characterized by the alteration of serum glycoproteins consisting of different monosaccharides. One of these is levo-fucose (L-fucose), a methyl pentose. Elevated levels of protein-bound fucose have been reported in various malignancies. Aim: The present study attempted to correlate levels of serum L-fucose as a biomarker with the various tumor node metastasis (TNM) stages of oral cancer. Methodology: The study was carried out on 90 subjects consisting of 30 healthy controls and 60 histopathologically proven oral squamous cell carcinoma (OSCC) cases. The serum fucose level estimation was done based on the method adopted by Winzler. Statistical analysis included independent sample's t-test, one-way ANOVA test, Karl–Pearson correlation test, and Tukey's HSD post hoc test to evaluate the significance and variability of values between groups. Results: Significant elevation in serum fucose levels was noticed among OSCC patients when compared with the controls and a progressive ascent of L-fucose levels were noted as the stage of severity increased. Serum fucose levels were independent of histopathological grading, age, and sex. Conclusion: Serum L-fucose levels were increased in OSCC patients, and a positive correlation was observed between serum L-fucose levels and TNM staging of OSCC. Thus, serum L-fucose can be used as an effective diagnostic and prognostic biomarker in OSCC patients.Hide Abstract
Megson, Z. A., Koerdt, A., Schuster, H., Ludwig, R., Janesch, B., Frey, A., Naylor, K., Wilson, I. B. H., Stafford, G. P., Messner, P. & Schäffer, C. (2015). Virulence, 6(3), 282-292.
The periodontal pathogen Tannerella forsythia expresses several glycosidases which are linked to specific growth requirements and are involved in the invasion of host tissues. α-L-Fucosyl residues are exposed on various host glycoconjugates and, thus, the α-L-fucosidases predicted in the T. forsythia ATCC 43037 genome could potentially serve roles in host-pathogen interactions. We describe the molecular cloning and characterization of the putative fucosidase TfFuc1 (encoded by the bfo_2737 = Tffuc1 gene), previously reported to be present in an outer membrane preparation. In terms of sequence, this 51-kDa protein is a member of the glycosyl hydrolase family GH29. Using an artificial substrate, p-nitrophenyl-α-fucose (KM 670 µM), the enzyme was determined to have a pH optimum of 9.0 and to be competitively inhibited by fucose and deoxyfuconojirimycin. TfFuc1 was shown here to be a unique α(1,2)-fucosidase that also possesses α(1,6) specificity on small unbranched substrates. It is active on mucin after sialidase-catalyzed removal of terminal sialic acid residues and also removes fucose from blood group H. Following knock-out of the Tffuc1 gene and analyzing biofilm formation and cell invasion/adhesion of the mutant in comparison to the wild-type, it is most likely that the enzyme does not act extracellularly. Biochemically interesting as the first fucosidase in T. forsythia to be characterized, the biological role of TfFuc1 may well be in the metabolism of short oligosaccharides in the periplasm, thereby indirectly contributing to the virulence of this organism. TfFuc1 is the first glycosyl hydrolase in the GH29 family reported to be a specific α(1,2)-fucosidase.Hide Abstract
Fan, S., Zhang, H., Chen, X., Lu, L., Xu, L. & Xiao, M. (2015). Journal of Basic Microbiology, 56(4), 347-357.
α-L-Fucosidases are key enzymes for the degradation of intestinal glycans by gut microbes. In this work, three putative α-L-fucosidases (Afc1, Afc2, and Afc3) genes from Clostridium perfringens ATCC 13124 were cloned and expressed in Escherichia coli. Afc1 had the α-L-fucosidase domain of glycoside hydrolase (GH) 29 family but showed no enzyme activity toward all the substrates examined. The putative acid/base residue of Afc1, Ser205, was replaced by a glutamic acid which is conserved in GH29-B α-L-fucosidases. However, the mutant Afc1-S205E still failed to show enzyme activity. Afc2 and Afc3 were determined to be 1,3-1,4-α-L-fucosidase of GH29-B subfamily and 1,2-α-L-fucosidase of GH95 family, respectively, and both of them could release fucose from porcine gastric mucin (PGM). When C. perfringens ATCC 13124 grew with the presence of PGM, the transcription of afc1 decreased slightly, while those of afc2 and afc3 increased to 2.2-fold and 1.4-fold, respectively, and the enzyme activities of Afc2 and Afc3 in the culture increased to 2.2-fold and 2.6-fold, respectively. These results suggest that Afc2 and Afc3 are involved in the degradation of intestinal fucosyl glycans by C. perfringens ATCC 13124.Hide Abstract