CD BioGlyco offers the 6'-siallylactose-based cell-free system construction service. This service enables the creation of a cell-free biosynthesis platform for the synthesis of a specific glycan structure 6'-siallylactose.

The Reliable and Custom 6'-Siallylactose-based Cell-free System Construction at CD BioGlyco

This service enables the synthesis of 6'-siallylactose using α2-6 sialyltransferase enzymes like HsSIAT1, PdST6, and PlST6, which transfer sialic acid to lactose's 6' position. The Cell-free Biosynthesis Platform allows the modular construction of protein glycosylation pathways. The system utilizes three plasmids: one for CMP-Sia synthesis containing the N. meningitidis CMP-Sia synthase gene, another for the target protein inducible with IPTG, and a third plasmid containing glycosyltransferases within the GT operon. This system successfully modifies the Fc region of engineered Fc protein with 6'-siallylactose glycans, confirmed by intact protein mass spectrometry analysis and exoglycosidase digestions.

CD BioGlyco offers a range of synthetic biology-based glycan production services. Leveraging our Cell Library, we select suitable cell expression systems according to the client's needs. We also provide the Development of Production Chassis Cells and subsequent Screening and Fermentation Engineering services. Additionally, we offer a portfolio of Pre-developed Glycan Products.

Fig.1 The process of construction of the 6'-siallylactose-based cell-free system. (CD BioGlyco)

• Prepare the cell-free extract: Start by preparing a cell lysate from Escherichia coli cells. The lysate can be obtained by growing E. coli cells, harvesting them, and then lysing them using a suitable method. The lysate should be kept on ice and then spun to collect the supernatant, which is used as the cell-free extract.
• Assemble the cell-free glycosylation reactions: Mix the cell-free extract with the necessary components for glycosylation reactions. These components include the target glycoprotein, glycosyltransferases, and activated sugar donors. For the synthesis of 6'-siallylactose, we would need the glycosyltransferase enzyme that adds sialic acid onto lactose in an α2,6 linkage.
• Optimize reaction conditions: Determine the optimal concentrations of substrates and cofactors for the glycosylation reactions.
• Incubate the reactions: Allow the glycosylation reactions to incubate overnight at an appropriate temperature.
• Purify the glycosylated product: After the incubation, isolate the glycosylated product using an affinity-tag purification method. This helps separate the desired glycosylated protein from other reaction components.
• Analyze the product: Confirm the presence of 6'-siallylactose on the glycoprotein by using mass spectrometry (MS) analysis.
• The specific details and conditions may vary depending on the specific glycosylation system and enzymes used. Confirm the successful glycosylation of the target proteins by analyzing them using techniques such as mass spectrometry or glycoproteomics. This verifies the functionality of the constructed cell-free system.

Applications

• The synthesized 6'-siallylactose can be incorporated into biopharmaceuticals to modulate their properties, such as stability, solubility, and immunogenicity.
• Researchers can use the 6'-siallylactose-based cell-free system to study the role of specific glycan structures in biological processes. This includes investigating glycan-protein interactions, cell signaling pathways, and host-pathogen interactions.
• The synthesized 6'-siallylactose serves as a reference standard or substrate for diagnostic assays, particularly those targeting sialic acid-binding proteins or lectins.

Advantages

• CD BioGlyco ensures the quality and reproducibility of synthesized glycans through rigorous quality control measures, including analytical testing and validation procedures, providing clients with confidence in the reliability of the produced glycan structures.
• The cell-free system construction service accelerates the glycan synthesis process by bypassing the need for cell culture and purification steps associated with traditional methods. This reduces both time and costs associated with glycan production.
• Our service allows for the customization of cell-free systems to meet specific research or industrial requirements. Clients can tailor the system components, such as glycosyltransferases and substrates, to their unique needs.

Similar Services at CD BioGlyco

At CD BioGlyco, our the 6'-siallylactose-based cell-free system construction service enables diverse applications in basic research, biopharmaceutical development, and diagnostic technologies, contributing to advancements in glycoscience and related fields. Please feel free to contact us for more information and ask for a custom quoetation.

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