Research Tool for Secretome Analysis — Azido Sugars

Currently, mass spectrometry-based proteomic analysis enables systematic research on secreted proteins [2]. However, this method cannot enrich secreted proteins, resulting in a small number of identified secreted proteins. In addition, conventional secretome analysis uses serum-free medium for cell culture to reduce sample complexity, and long-term serum-free culture is likely to alter cell activity.

(Panel a: Chemical structures of N-Glycans, O-Glycans with azido groups (N3), including Sialic acid, Galactose, GlcNAc, Mannose, GalNAc, Fucose, and azido sugar analogs such as GlcNAz, ManNAz, GalNAz; Panel b: Schematic diagram of metabolic labeling and enrichment process: Azido sugars are metabolically labeled into secreted proteins in cells, then covalently labeled with biotin via click chemistry; after trypsin digestion and PNGase F treatment, deglycosylated peptides are analyzed by LC-MS)

In recent years, unnatural sugars containing bioorthogonal groups (such as azido groups) have been used for metabolic labeling of glycosylated proteins, thereby enabling cellular imaging of glycoproteins or selective enrichment for proteomic analysis.

This strategy consists of two steps: Azido sugar analogs (Figure 1a) are added to the cell culture medium and incorporated into glycoproteins through intracellular glycan biosynthesis pathways; then, covalent labeling with imaging probes or affinity probes is performed specifically via click chemistry (Figure 1b). Since secreted proteins are usually glycoproteins, this metabolic glycan labeling has been used for the labeling and enrichment of secreted proteins. N-azidoacetylgalactosamine (GalNAz), N-azidoacetylglucosamine (GlcNAz), and N-azidoacetylmannosamine (ManNAz) are the most classic azido sugar analogs, and their applications are shown in Table 1.

Table 1 Several Commonly Used Azido Sugar Metabolic Probes [4-6]

With the development of technology, the above three most commonly used commercial unnatural sugar metabolic labeling products (Table 1) have evolved into Ac4ManNAz, Ac4GlcNAz, and Ac4GalNAz, which are tetraacylated N-azidoacetylmannosamine, N-azidoacetylglucosamine, and N-azidoacetylgalactosamine, respectively. Acylation increases their solubility in many solvents, making the handling of these reagents easier.

Features of Azido Sugars

•         Bioorthogonality — The azido group is small and non-reactive, and does not exist in living systems. Therefore, azido-sugar compounds will not interfere with endogenous cellular pathways or replace their naturally occurring analogs;

•         Compatibility — Phosphorus compounds can be effectively used for reaction chemistry under simple buffer conditions, without the need for auxiliary reagents such as copper or reducing agents;

•         Chemical Selectivity — Azides and phosphine groups do not react with or interfere with components of biological samples, but couple with each other efficiently;

•         Versatility — Azide labeling can be used for detection, immobilization, conjugation, or affinity purification, depending on the phosphine-activated compound it reacts with.

These sugars are azide derivatives of natural monosaccharides, and cells use post-translational modification biochemical pathways to glycosylate proteins. The azide functional group is small and does not react with endogenous molecules. When delivered to cells, these compounds are integrated into glycosylation events to effectively "label" glycoproteins with azide groups. The azide groups can then be specifically used for detection or conjugation using alkyne-activated reagents ("click" chemistry) or phosphine-activated reagents (Staudinger ligation).

When used in combination with phosphine-activated fluorescent dyes, biotin reagents, or other compounds, these azide-modified sugars facilitate the study of cellular pathways involving glycosylation.

Product Information

Brand Mission

ANT BIO PTE. LTD. is dedicated to advancing life science research by providing high-quality, reliable reagents and comprehensive solutions. We recognize the critical challenges in secretome research and the urgent need for standardized, specialized experimental tools. Through our specialized sub-brands (Absin, Starter, UA), we have developed a targeted product portfolio for secretome and glycosylation research, covering core azido sugar metabolic probes (GlcNAz, GalNAz, ManNAz, and their tetraacylated derivatives) and related supporting reagents.

Our team adheres to stringent quality control standards throughout the product development and production process, ensuring the high purity, stability, and biological activity of each product. We are committed to providing professional technical support and customer-centric services, helping researchers overcome experimental challenges such as low enrichment efficiency of secreted proteins, altered cell activity in serum-free culture, and difficult operation of metabolic labeling reagents, and accelerating the pace of scientific research breakthroughs in secretome and glycosylation-related fields. ANT BIO PTE. LTD. strives to be a trusted partner for scientists worldwide, contributing to the advancement of proteomic research technology and the development of innovative biological detection methods.

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This article is AI-compiled and interpreted based on the original work related to azido sugars and secretome research. All intellectual property (e.g., experimental protocols, data, images) of the original publication shall belong to the relevant research team. For any infringement, please contact us promptly and we will take immediate action.

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ANT BIO PTE. LTD. – Empowering Scientific Breakthroughs

At ANTBIO, we are committed to advancing life science research through high-quality, reliable reagents and comprehensive solutions. Our specialized sub-brands (Absin, Starter, UA) cover a full spectrum of secretome and glycosylation research needs, from core azido sugar metabolic probes to related supporting products. With a focus on innovation, quality, and customer-centricity, we strive to be your trusted partner in overcoming secretome research challenges and driving progress in proteomic research. Explore our product portfolio today and elevate your research to new heights.