Cell-free glycoprotein synthesis platform enables creation of potent glycomedicines in a single pot with potential for small batch, decentralized biomanufacturing
Research led by PhD candidate Thapakorn Jaroentomeechai and former DLRG undergraduate Jessica Stark (CBE ’12), now a Ph.D. student in the Jewett group at Northwestern, are co-first authors on a newly published paper in Nature Communications. The article, “Single-pot glycoprotein biosynthesis using a cell-free transcription-translation system enriched with glycosylation machinery,” was the result of an NSF- and DTRA-funded collaboration between the DeLisa lab and Jewett Lab to develop a platform for cell-free synthesis of glycoprotein therapeutics and vaccines.
The emerging discipline of bacterial glycoengineering has made it possible to produce designer glycans and glycoconjugates for use as vaccines and therapeutics. Unfortunately, cell-based production of homogeneous glycoproteins remains a significant challenge due to cell viability constraints and the inability to control glycosylation components at precise ratios in vivo. To address these challenges, our novel cell-free glycoprotein synthesis (CFGpS) technology seamlessly integrates protein biosynthesis with asparagine-linked protein glycosylation. CFGpS leverages a glyco-optimized Escherichia coli strain to source cell extracts that are selectively enriched with glycosylation components, including oligosaccharyltransferases (OSTs) and lipid-linked oligosaccharides (LLOs). The resulting extracts enable a one-pot reaction scheme for efficient and site-specific glycosylation of target proteins. The CFGpS platform is highly modular, allowing the use of multiple distinct OSTs and structurally diverse LLOs. As such, we anticipate CFGpS will facilitate fundamental understanding in glycoscience and make possible applications in on-demand biomanufacturing of glycoproteins.
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