TECHNOLOGY BRIEF

CIS-ACTING PEPTIDE CHAPERONES

For further information, contact Christine Brakel

Description: The present invention provides a method for increasing the yield of soluble, properly folded recombinant proteins. Expressing proteins fused to short peptide extensions causes the protein to become either self-chaperoning in vivo or more recoverable from inclusion bodies in vitro.

Commercial, Technical and Competitive Merit: Advances in recombinant protein expression have resulted in the production of a wide variety and substantial quantities of valuable proteins for use in research and therapeutic and industrial applications. However, a commonly encountered obstacle to the production of useful recombinant proteins has been that many times the over-expression of the recombinant protein results in an incorrectly folded protein that aggregates into an insoluble entity commonly referred to as an inclusion body. While many attempts to overcome this aggregation problem have been proposed, few solutions, if any, are applicable across a wide variety of proteins.

The present invention includes materials and methods for production of proteins fused to short peptide extensions that enhance the ability of the proteins to attain their native conformations. In some cases, the peptide extensions caused proteins, which had previously formed insoluble inclusion bodies, to be over-expressed as completely soluble, properly folded proteins. In other cases, solubilization by expression with the peptide extension was only partially successful. In these circumstances the peptide extensions enhanced the recovery of properly folded protein following chemical and heat denaturation and renaturation of the protein in vitro. Proteins could be renatured from inclusion bodies at concentrations exceeding 5mg/ml. Furthermore, brief protease treatments of the soluble fusion proteins cleaved the peptide extensions from the proteins of interest. Antibodies to the peptide extensions provide a simplified means of isolating the expressed fusion proteins or monitoring the proteolytic removal of the peptide extension.

Applications:

• It is expected that the chaperoning peptide extensions will be effective in any expression system in which inclusion body formation is a problem.
• The peptide extensions eliminate the need to alter to culture conditions to reduce the extent or likelihood of inclusion body formation.
• The peptide extensions are useful for recovery of proteins in vitro from inclusion bodies that continue to persist despite expression in fusion with the extension.
• Properties of bacterial cell biocatalysts containing recombinant proteins with a propensity to form inclusion bodies can be enhanced by the methods of this invention.

Inventors: Paul I. Freimuth, Yian-Biao Zhang and Jason Howitt

Patent Status: U.S. Patent Application Serial No. 10/037,243 pending.

License Status: Available on a negotiable basis for research and commercial use.