Glycosaminoglycan Degradation Analysis Service

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Overview

The human gut microbiota (HGM) is a large microbial community that is vital for human health. HGM maintenance depends on its ability to utilize complex polysaccharides in the form of dietary and host polysaccharides as a nutritional source. The mechanism by which the community metabolizes these polysaccharides is therefore essential to understanding the ecology of the system and how it can be manipulated to benefit human health. Bacteroidetes are one of the two major phylum that control HGM. Up to 20% of the genome of these organisms is dedicated to complex glycan metabolism in the form of genes that mainly encode carbohydrate active enzymes (e.g., polysaccharide lyase (PLs), glycosidase, glycan transport and binding proteins, and carbohydrate sulfatase).

The human gut microbiota (HGM), which is essential for human health, utilizes complex polysaccharides as its main carbon source. Glycosaminoglycans are an important and highly preferred source of nutrition for HGM. The metabolic pathways of various glycosaminoglycan substrates remain unknown. There have been researches on the biochemical, genetic and structural anatomy of its genetic locus, the glycosaminoglycan arrangement in organisms metabolizing Bacteroides polymorpha. It was pointed out that two previously unknown surface glycan-binding proteins were found that promoted the introduction of glycosaminoglycans into the periplasm. The metabolic pathways of glycosaminoglycans are determined by different kinetics and genetic specificities of various periplasmic lyases.

Creative Proteomics, as the leader of bio-tech industry, devote to forge high quality targeted metabolomics service with advanced apparatus and professional research team. We provide high throughput, high precision and short time-consuming Glycosaminoglycan Degradation Analysis Service, which could be meet all the need of our customers, to assist your scientific research.

Advantages

Short time-consuming
High sensitivity and few detection restrictions
High precision and good repeatability
High throughput
Customized service

Fig 1. Model of glycosaminoglycan degradation by PUL CS/DS/HA.

Service workflow

Extract and purifying glycosphingolipid from cell membrane or serum
Release that glycosphingolipid glycan with the glucosidase
The total methylation of glycan
UPLC-HILIC-FLD analysis based on 2-AB labeling
Analysis of fully methylated glycans by time-of-flight mass spectrometry

Fig 2. Workflow for the discovery-based proteomics.

Sample requirement

Serum/plasma: 500 μl/sample

Repeated freezing and thawing of samples must be avoided. The serum sample should be precipitated in the collection tube for 30 minutes at room temperature, then transported to the centrifuge tube and centrifuged at 8000 rpm for 5 minutes. After centrifugation, the supernatant was equally divided into a freezing tube of 500 uL / sample.

Protein: 100 µg
Anticoagulated blood (EDTA): 1 mL

Anticoagulants and preservatives must be added immediately after collection and then frozen at -80 °C.

Urine: 1 ml/sample

Urine samples should be equally divided into centrifuge tubes with 1 mL per tube, each tube is added with 1/100 (w/v) sodium azide and stored at -80 °C.

Animal tissues: 200 mg/sample

Samples should be frozen in liquid nitrogen immediately and then transported to -80 °C for storage after collected.

Cells: ≥ 1 × 10⁷/sample

Cytoactive should be terminated immediately to maintaining cell integrity.

Feces: 500 mg/sample

In general, to assure enough sample to fulfill the whole project, the volume of the single sample need to be offered as much as possible. The remaining samples will be stored for one year free of charge and returned at any time if necessary. All samples need to be stored and transported at -80°C and try to avoid using surfactants (SDS, Triton-X) and inorganic salts.

Clinical samples are repeated in no less than 30 cases in a single group.

Animal samples are repeated in no less than 9 cases in a single group.

Report delivery

Experimental procedure
Parameters of HPLC and MS
Raw data, chromatograms and mass spectra
Metabolites quantification data
Custom analysis report

Service cycle

Sample testing: 5-10 working days
Data analysis: 5-10 working days

Creative Proteomics metabolism analysis platform is committed to the all-around, reliable and accurate analysis service for a variety of target substances, which is suitable for life-science research, drug exploration, biological determination and other fields. We sincerely hope to cooperate with you to assistant your scientific research.

References

J.A.D. Van, S. Clotet-Freixas, A.C. Hauschild, et al. Urinary proteomics links keratan sulfate degradation and lysosomal enzymes to early type 1 diabetes. PLoS One, 2020, 15(5):e0233639.
W. Wang, L. Shi, Y. Qin, et al. Research and Application of Chondroitin Sulfate/Dermatan Sulfate-Degrading Enzymes. Front Cell Dev Biol, 2020, 8:560442.
Z. Wang, K. Arnold, Y. Xu, et al. Quantitative analysis of heparan sulfate using isotopically labeled calibrants. Commun Biol, 2020, 3(1):425.