Scalable VHH and Peptide Secretion Platform Using C. glutamicum

I. E. coli Expression: A Standard Choice for Protein Production

E. coli is a microbial expression system commonly used to manufacture proteins. It remains popular due to its growth speed, simple upstream, low cost and relative scalability. In terms of challenges, standard E. coli accumulates proteins inside the cell, which often form insoluble inclusion bodies that require cell harvesting and disruption, inclusion body collection and solubilization, protein refolding and complex purification. Another issue is high endotoxin levels since E. coli is a gram-negative bacterium. Endotoxin removal requires a downstream process in which the yield can be very low for certain proteins. E. coli might also struggle to make peptides due to proteolytic activities inside the cell. E. coli is a widely used platform with advantages, but its challenges should also be considered when determining whether it is the best method for specific proteins.

II. Solid-Phase Synthesis: A Small-Scale Production Method for Short Peptides

Solid-phase synthesis is a popular method for quickly manufacturing small volumes of peptides, since E. coli and/or other fermentation methods often require peptide-specific development work upfront. Solid-phase synthesis is also practical for incorporating non-natural amino acids or other modifications commonly used in the manufacturing of pharmaceutical peptides. Whether solid-phase synthesis is the best method depends on the structure, development phase and volumes/scale of the peptide of choice. Making peptides at a large scale via solid-phase synthesis requires large amounts of hazardous reagents and solvents, and economies of scale are limited. Despite these challenges, solid-phase synthesis remains an excellent choice for small-scale peptide production, and various technologies are emerging to address its drawbacks.

III.  A Productive, Scalable and Sustainable Platform to Secrete VHH and Peptides

An alternative microbial secretion platform based on Corynebacterium glutamicum offers a productive, scalable and sustainable method for manufacturing VHHs, peptides and other biologics. This gram-positive, non-sporulating and non-endotoxic bacterium has a long history of use in industrial applications. The platform can secrete soluble, correctly folded and biologically active proteins into the culture media with high titers and purity without endotoxins, drastically simplifying the downstream process. Another advantage is that it does not secrete proteases, enabling recovery of intact target molecules, including peptides, which are often susceptible to degradation via proteases. A significant reduction in solvent use and Process Mass Intensity (PMI), in addition to superior scalability and economies of scale, enables effective scaling up from solid-phase synthesis for peptides.

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