UBIQUITIN FUSION TECHNOLOGY - BIOPROCESSING OF PEPTIDES

Ubiquitin fusion technology represents an emerging method for economic ally producing peptides and small proteins in the bacterium Escherichi a coli. Our focus is on peptide production where the need for cost-eff ective, scaleable processes has recently been highlighted by Kelley (1 996). There are two principal features: (1) the expression system cons ists of a suitable E. coli host strain paired with a plasmid that enco des the ubiquitin fusion and (2) an ubiquitin-specific protease, UCH-L 3, which cleaves only C-terminal extensions from ubiquitin. In this wo rk, multigram yields were obtained of four ubiquitin fusions derived f rom cell paste generated in single 10-L fermentations. All were expres sed intracellularly and remained soluble at extremely high levels of e xpression. Bacterial freeze-thaw lysates contained over 95% pure ubiqu itin fusion protein. All four fusions were efficiently cleaved to ubiq uitin and the peptide products. In one case, the final yield of peptid e was 1.08 g from 3 L of low cell density bacterial culture. The combi nation of exceptional overexpression of the ubiqutin-peptide fusion pr oteins and a robust and specific protease are unique advantages contri buting to a cost-effective, scaleable, and generic bioprocess for pept ide production.