Bacterial production of biologically active canine interleukin-1β by seamless SUMO tagging and removal

• Published in: Protein expression and purification 2006-11, Vol.50 (1), p.102-110
• Main Authors: Kirkpatrick, Robert B., Grooms, Michael, Wang, Feilan, Fenderson, Heather, Feild, John, Pratta, Michael A., Volker, Craig, Scott, Gilbert, Johanson, Kyung
• Format: Article
• Language: English
• Subjects: Canine interleukin 1β; Escherichia coli; Osteoarthritis; Seamless cloning; Small ubiquitin related modifier (SUMO)
• ISSN: 1046-5928; 1096-0279
• DOI: 10.1016/j.pep.2006.05.022

Interleukin 1β (IL-1β) is a potent stimulator of extracellular matrix degradation in models of osteoarthritis (OA). In contrast to bovine explant models which effectively respond to recombinant human IL-1β, canine models are relatively refractory to human IL-1β stimulation. Canine IL-1β cDNA was cloned in order to produce a fully potent species matched preparation of IL-1β for use specifically in canine models of OA. Established methods for the production of various orthologous IL-1β proteins from different species are problematic due to the exquisite sensitivity of the mature IL-1β product to N-terminal variations and the intrinsic technical challenges associated with producing an unmodified product. We have applied a seamless method of SUMO tagging and removal in order to produce a homogeneous unmodified preparation of canine IL-1β from Escherichia coli which was found to be a potent inducer of aggrecanase activity in isolated canine articular chondroctyes. This method combines highly efficient aspects of seamless plasmid engineering, protein purification, and precise tag removal.