The Custom 2'-Fucosyllactose-based Cell-free System Construction at CD BioGlyco

CD BioGlyco's Cell-Free Platform revolutionizes the assembly of protein glycosylation pathways, eliminating the need for cloning and transformation in Living Cells. A pivotal component of our service involves integrating N-glycosyltransferases (NGTs), soluble enzymes proficient in transferring glucose residues onto proteins within Escherichia coli's cytoplasm. Combined with co-expressed glycosyltransferases (GTs), these NGTs present a promising solution for synthesizing intricate glycoproteins typically challenging to produce via conventional methods.

Fig.1 The process of 2'-fucosyllactose-based cell-free system construction. (CD BioGlyco)

• Identify and select the enzymes involved in 2'-fucosyllactose biosynthesis, including those for fucose activation and lactose fucosylation.
• Express and purify the chosen enzymes using recombinant DNA methods in a suitable host like E. coli, followed by standard protein purification procedures.
• Prepare the necessary substrates, such as lactose and fucose, required for the enzymatic reactions.
• Optimize reaction conditions like pH, temperature, and enzyme concentrations for each enzymatic step.
• Design and optimize the coupling of enzymatic reactions to ensure efficient lactose conversion to 2'-fucosyllactose.
• Develop strategies for cofactor regeneration, crucial for certain enzymatic steps necessitating cofactors like NADH or ATP.
• Establish methods for monitoring reaction progress and quantifying 2'-fucosyllactose production, employing analytical techniques like HPLC or MS.
• Scale up the optimized cell-free system to larger volumes, employing larger vessels or continuous flow systems.
• Devise purification methods to isolate and purify the synthesized 2'-fucosyllactose from the reaction mixture, utilizing techniques like chromatography or membrane filtration.
• Characterize the purified 2'-fucosyllactose to verify its identity and purity, employing techniques such as NMR spectroscopy or MS.

Applications of 2'-Fucosyllactose-based Cell-free System

• Our 2'-fucosyllactose-based cell-free system production eliminates the need for maintaining and culturing living cells, reducing the cost associated with cell culture media, equipment, and labor.
• Our cell-free systems can achieve higher production rates compared to living cells. The absence of cellular growth and maintenance processes allows for the efficient conversion of substrates into the desired product.
• Our cell-free systems can be easily optimized and engineered to enhance the production of specific compounds. This flexibility allows for rapid screening and optimization of reaction conditions, leading to improved production efficiency.

Advantages

• CD BioGlyco prioritizes cost-effectiveness in research and development endeavors, offering solutions that optimize resources while upholding quality and performance standards.
• Stringent quality control measures are maintained by CD BioGlyco throughout the construction process of the 2'-fucosyllactose-based cell-free system, ensuring the reliability and reproducibility of systems provided to clients.
• CD BioGlyco is dedicated to timely project delivery, supported by efficient project management practices. Clients can expect prompt delivery of our 2'-fucosyllactose-based cell-free systems, facilitating seamless progress in their research endeavors.

Similar Services at CD BioGlyco

At CD BioGlyco, our researchers synthesize and assemble three-enzyme biosynthetic pathways for the production of fucosylated lactose structures. By screening and selecting appropriate GTs, including α1,3 and α1,2 fucosyltransferases, clients can customize glycosylation pathways tailored to their specific needs. Besides, CD BioGlyco provides Chassis Development services for clients to develop suitable production cells, also clients can find our Synthetic Biology-associated Products on our website. Please feel free to contact us if you are interested in our services.

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