Elastin-like polypeptides as a purification tag for recombinant proteins.
This unit presents a recombinant protein purification method that employs an elastin-like polypeptide (ELP) as a purification tag. ELPs undergo a sharp and reversible phase transition when heated above their lower critical solution temperature. ELPs retain this behavior when they are fused to a protein, and thereby provide a simple method to isolate a recombinant ELP fusion protein from cell contaminants by cycling the solution through the insoluble and soluble phase of the ELP fusion protein using a procedure that is termed Inverse Transition Cycling. This method does not require the use of chromatography, so it is cost-effective, easy to scale up, and easy to multiplex.
Hassouneh
Wafa
W
Duke University, Durham, North Carolina.
Christensen
Trine
T
Duke University, Durham, North Carolina.
Chilkoti
Ashutosh
A
Duke University, Durham, North Carolina.
D002846:Chromatography, Affinity; D004549:Elastin / Q000737:chemistry* / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D010316:Particle Size; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000302:isolation & purification*; D013696:Temperature
D010455:Peptides; D011993:Recombinant Fusion Proteins; D004549:Elastin
Banki, M.R., Feng, L.A., and Wood, D.W. 2005. Simple bioseparations using self-cleaving elastin-like polypeptide tags. Nat. Methods 2:659-661.
Chen, T.H.H., Bae, Y., and Furgeson, D.Y. 2008. Intelligent biosynthetic nanobiomaterials (IBNs) for hyperthermic gene delivery. Pharm. Res. 25:683-691.
Cho, Y.H., Zhang, Y.J., Christensen, T., Sagle, L.B., Chilkoti, A., and Cremer, P.S. 2008. Effects of Hofmeister anions on the phase transition temperature of elastin-like polypeptides. J. Phys. Chem. B 112:13765-13771.
Christensen, T., Trabbic-Carlson, K., Liu, W.G., and Chilkoti, A. 2007. Purification of recombinant proteins from Escherichia coli at low expression levels by inverse transition cycling. Anal. Biochem. 360:166-168.
Christensen, T., Amiram, M., Dagher, S., Trabbic-Carlson, K., Shamji, M., Setton, L., and Chilkoti, A. 2009. Fusion order controls expression level and activity of elastin-like polypeptide fusion proteins. Protein Sci. 18:1377-1387.
Farewell, A. and Neidhardt, F.C. 1998. Effect of temperature on in vivo protein synthetic capacity in Escherichia coli. J. Bacteriol. 180:4704-4710.
Fujita, Y., Funabashi, H., Mie, M., and Kobatake, E. 2007. Design of a thermocontrollable protein complex. Bioconjug. Chem. 18:1619-1624.
Ge, X., Trabbic-Carlson, K., Chilkoti, A., and Filipe, C.D.M. 2006. Purification of an elastin-like fusion protein by microfiltration. Biotechnol. Bioeng. 95:424-432.
Hofmeister, F. 1888. Zur Lehre von der Wirkung der Salze. Naunyn Schmiedebergs Arch. Pharmacol. 24:247-260.
Jenikova, G., Lao, U.L., Gao, D., Mulchandani, A., and Chen, W. 2007. Elastin-calmodulin scaffold for protein microarray fabrication. Langmuir 23:2277-2279.
Kim, B., and Chilkoti, A. 2008. Allosteric actuation of inverse phase transition of a stimulus-responsive fusion polypeptide by ligand binding. J. Am. Chem. Soc. 130:17867-17873.
Kim, J.Y., Mulchandani, A., and Chen, W. 2005. Temperature-triggered purification of antibodies. Biotechnol. Bioeng. 90:373-379.
Lao, U.L., Kostal, J., Mulchandani, A., and Chen, W. 2007. Affinity purification of plasmid DNA by temperature-triggered precipitation. Nat. Protoc. 2:1263-1268.
Luan, C.H., Harris, R.D., Prasad, K.U., and Urry, D.W. 1990. Differential scanning calorimetry studies of the inverse temperature transition of the polypentapeptide of elastin and its analogs. Biopolymers 29:1699-1706.
Massodi, I., and Raucher, D. 2007. A thermally responsive Tat-elastin-like polypeptide fusion protein induces membrane leakage, apoptosis, and cell death in human breast cancer cells. J. Drug Target. 15:611-622.
McPherson, D.T., Xu, J., and Urry, D.W. 1996. Product purification by reversible phase transition following Escherichia coli expression of genes encoding up to 251 repeats of the elastomeric pentapeptide GVGVP. Protein Expr. Purif. 7:51-57.
Meyer, D.E. and Chilkoti, A. 1999. Purification of recombinant proteins by fusion with thermally-responsive polypeptides. Nat. Biotechnol. 17:1112-1115.
Meyer, D.E. and Chilkoti, A. 2002. Genetically encoded synthesis of protein-based polymers with precisely specified molecular weight and sequence by recursive directional ligation: Examples from the elastin-like polypeptide system. Biomacromolecules 3:357-367.
Meyer, D.E. and Chilkoti, A. 2004. Quantification of the effects of chain length and concentration on the thermal behavior of elastin-like polypeptides. Biomacromolecules 5:846-851.
Meyer, D.E., Trabbic-Carlson, K., and Chilkoti, A. 2001. Protein purification by fusion with an environmentally responsive elastin-like polypeptide: Effect of polypeptide length on the purification of thioredoxin. Biotechnol. Prog. 17:720-728.
Shamji, M.F., Betre, H., Kraus, V.B., Chen, J., Chilkoti, A., Pichika, R., Masuda, K., and Setton, L.A. 2007. Development and characterization of a fusion protein between thermally responsive elastin-like polypeptide and interleukin-1 receptor antagonist: Sustained release of a local antiinflammatory therapeutic. Arthritis Rheum. 56:3650-3661.
Shimazu, M., Mulchandani, A., and Chen, W. 2002. Environmentally triggered purification and immobilization of Elastin-OPH fusions. Abstr. Pap. Am. Chem. Soc. 224:U228-U228.
Stano, N.M. and Patel, S.S. 2004. T7 lysozyme represses T7 RNA polymerase transcription by destabilizing the open complex during initiation. J. Biol. Chem. 279:16136-16143.
Trabbic-Carlson, K., Liu, L., Kim, B., and Chilkoti, A. 2004a. Expression and purification of recombinant proteins from Escherichia coli: Comparison of an elastin-like polypeptide fusion with an oligohistidine fusion. Protein Sci. 13:3274-3284.
Trabbic-Carlson, K., Meyer, D.E., Liu, L., Piervincenzi, R., Nath, N., LaBean, T., and Chilkoti, A. 2004b. Effect of protein fusion on the transition temperature of an environmentally responsive elastin-like polypeptide: A role for surface hydrophobicity? Protein Eng. Des. Sel. 17:57-66.
Wu, W.Y., Mee, C., Califano, F., Banki, R., and Wood, D.W. 2006. Recombinant protein purification by self-cleaving aggregation tag. Nat. Protoc. 1:2257-2262.
Zhang, Y.J. and Cremer, P.S. 2006. Interactions between macromolecules and ions: The Hofmeister series. Curr. Opin. Chem. Biol. 10:658-663.