CD BioGlyco provides a bespoke service for developing N-linked 3'-siallylactose-based cell-free systems. This comprehensive service encompasses the design, assembly, and fine-tuning of cell-free systems customized for N-linked 3'-siallylactose production.

The Custom Cell-free System for N-linked 3'-Siallylactose Production at CD BioGlyco

CD BioGlyco presents a tailored service for constructing N-linked 3'-siallylactose-based cell-free systems, employing a novel modular, two-pot method for swift assembly and screening of synthetic glycosylation pathways in vitro. This method involves selectively enriching crude E. coli lysates with individual glycosyltransferases (GTs) using cell-free protein synthesis (CFPS) expression, which are then combined to form multi-enzyme glycosylation pathways. Leveraging N-glycosyltransferases (NGTs), the approach bypasses substrate specificity constraints of oligosaccharyltransferases (OSTs) and obviates the need for protein transport across membranes or lipid-associated components. This technology enhances glycoprotein synthesis efficiency and versatility, streamlining glycosylation pathway development for various applications in glycobiology, biotechnology, and pharmaceuticals.

Using our extensive Cell Library, CD BioGlyco offers diverse chassis for glycan production development, complemented by custom Strain Development services to meet individual client requirements.

Our standard procedure for constructing an N-linked 3'-siallylactose-based cell-free system is outlined below.

Fig.1 The process of construction of N-linked 3'-siallylactose-based cell-free system. (CD BioGlyco)

• Design plasmids encoding the necessary enzymes for the biosynthesis of N-linked 3'-siallylactose. This includes the NGT enzyme, the lactose synthase enzyme, and the sialyltransferase enzyme.
• Clone the genes encoding the NGT, lactose synthase, and sialyltransferase enzymes into separate plasmids. Use standard molecular biology techniques such as PCR, restriction enzyme digestion, and ligation to insert the genes into the plasmids.
• Prepare a crude lysate from E. coli cells. Initially, cells are lysed and cellular debris is removed, yielding a lysate containing the essential cellular components for protein synthesis.
• Perform CFPS using the prepared lysate and the plasmids containing the genes for the NGT, lactose synthase, and sialyltransferase enzymes. This allows the enzymes to be produced in vitro.
• Next, the lysates containing the expressed enzymes are mixed and matched to construct the multi-enzyme glycosylation pathway. This allows the enzymes to work together to synthesize N-linked 3'-siallylactose on target proteins.
• 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

• Researchers can utilize the constructed cell-free systems to engineer glycoproteins with precise N-linked 3'-siallylactose modifications.
• The cell-free systems can be employed in drug discovery efforts, particularly in screening assays for identifying glycoproteins with specific binding properties to targets of interest.
• The ability to synthesize N-linked 3'-siallylactose-modified glycoproteins in a cell-free system is valuable for elucidating the role of glycosylation in biological processes.
• The construction of N-linked 3'-siallylactose-based cell-free systems enables the scalable production of complex glycoproteins for industrial applications.
• The cell-free systems offer a valuable educational tool for teaching and training purposes in academic settings.

Advantages

• CD BioGlyco offers customizable solutions tailored to the specific needs and requirements of each client. Whether it's glycan synthesis, glycoprotein modification, or glycoanalysis, we work closely with clients to develop personalized strategies and deliver optimal results.
• CD BioGlyco offers a comprehensive range of glycan-related services, including synthesis, modification, analysis, and purification.
• CD BioGlyco is dedicated to advancing the field of glycobiology through continuous innovation and research.

Similar Services at CD BioGlyco

At CD BioGlyco, our N-linked 3'-siallylactose-based cell-free system construction service offers a reliable and efficient solution for researchers and biotech companies seeking to produce N-linked 3'-siallylactose for various applications in glycobiology, biotechnology, and pharmaceuticals. And we also offer diverse Glycan Products based on our synthetic biology associated production lab. Please feel free to contact us if you are interested in our service and would like to know more about us.

• 6'-Siallylactose-based Cell-free System Construction Service
• αGal Epitope Synthesis-based Cell-free System Construction Service
• Pyruvylated Lactose-based Cell-free System Construction Service
• 2'-Fucosyllactose-based Cell-free System Construction Service
• 3-Fucosyllactose-based Cell-free System Construction Service
• Mono- or Di- Fucosylated Lactose-based Cell-free System Construction Service
• Mono- or Di-Fucosylated LacNAc-based Cell-free System Construction Service
• Sialylated Lactose-based Cell-free System Construction Service
• Sialylated LacNAc-based Cell-free System Construction Service