Princeton Invention #
Princeton University have developed a new economical method for the purification
of recombinant proteins expressed in E. coli. In this method, the host
cells simultaneously produce both an affinity tagged product protein and an
easily recovered granular affinity carrier. The self-cleaving capability of the
affinity tag allows the simple recovery of a native target protein at reasonable
yield and very low cost. This simple technique is likely to be portable to
numerous other expression hosts and is a significant advance in the large-scale
production of recombinant protein products. This system has been
successfully used at laboratory scale to purify several active test proteins at
reasonable yield. It is expected that this combination of improved economics and
simplicity will constitute a significant breakthrough in both large-scale
production of purified proteins and enzymes and high-throughput proteomic
studies of peptide libraries.
developed within the Chemical Engineering Department of Princeton University,
combines two well established technologies to generate a breakthrough in protein
production and purification. The first is the production of polyhydroxybutyrate
(PHB) granules in engineered strains of E. coli. The second is a recently
developed group of self-cleaving affinity tags based on protein splicing
elements known as inteins. By combing these technologies with a PHB-binding
protein, a self-contained protein expression and purification system has been
developed. In this system, the PHB-binding protein effectively acts as an
affinity tag for desired product proteins. The tagged product proteins are
expressed in an E. coli strain that also produces intracellular PHB
granules, where they bind to the granules via the PHB-binding tag. The granules
and attached proteins can then be easily recovered following cell lysis by
simple mechanical means such as filtration or centrifugation. Once purified, the
product protein is self-cleaved from the granules and released into solution in
a substantially purified form. By allowing the bacterial cells to effectively
produce both the affinity resin and tagged target protein, the cost associated
with the purification of recombinant proteins could be greatly reduced.
Gillies, A. R., Hsii, J. F., Oak, S.
& Wood, D. W., 2008, Rapid cloning and purification of proteins: Gateway
vectors for protein purification by self-cleaving tags (Editors¿ Choice feature
publication), Biotechnology and Bioengineering, Vol. 101 (2),
Mee, C., Banki, M. R. and Wood, D. W.,
2008, Towards the Elimination of Chromatography in Protein Purification:
Expressing Proteins Engineered to Purify Themselves, Chemical Engineering
Journal, Vol. 135, 56-62.
Banki, M.R. & Wood, D.W., 2005,
Inteins and Affinity Resin Substitutes for Protein Purification and Scale Up,
Microbial Cell Factories, Vol. 4:32.
Barnard, G.C., McCool, J.D., Wood,
D.W. & Gerngross T.U., 2005, An Integrated Recombinant Protein Expression
and Purification Platform, Applied and Environmental Microbiology, Vol. 71,
Banki,M., Gerngross,T., Wood,
D., 2005, Novel and economical purification of recombinant proteins:
Intein-mediated protein purification using in vivo polyhydroxybutyrate (PHB)
matrix association, Protein Science, Vol. 14, 1387-1395.
Princeton is currently seeking
industrial collaborators to commercialize this technology.
For more information on Princeton
University Invention # 04-2076 please contact:
Office of Technology Licensing and Intellectual Property
4 New South Building
Princeton, NJ 08544-0036
(609) 258-1159 fax