Abstract
Fusion protein technology has been creatively applied to solve many problems encountered in the study of protein structure and function (1,2). One prevalent application is the use of fusion proteins to improve protein expression in Escherichia coli and provide convenient methods for affinity-based protein purification. In many instances, when a foreign protein is overexpressed at high levels in E. coli, the majority of the protein is present as insoluble inclusion bodies. Previous research experience with fusion proteins containing glutathione S-transferase (GST) (3), maltose binding protein (MBP) (4), and thioredoxin (5) has shown that these proteins can improve the expression and solubility of many heterologous proteins. However, improvements in protein expression and solubility are not always guaranteed with these systems. The purpose of our recent research (6) has been to use a combination of protein solubility modeling, bioinformatics, and molecular biology techniques to systematically identify native E. coli proteins which have maximal potential for increasing recombinant fusion protein solubility.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
LaVallie, E. R. and McCoy, J. M. (1995) Gene fusion expression systems in Escherichia coli. Curr. Opin. Biotechnol. 6, 501–506.
Uhlen, M., Forsberg, G., Moks, T., Hartmanis, M., and Nilsson, B. (1992) Fusion proteins in biotechnology. Curr. Opin. Biotechnol. 3, 363–369.
Smith, D. B. and Johnson, K. S. (1988) Single-step purification of polypeptides expressed in Escherichia coli as fusions with glutathione S-transferase. Gene 67, 31–40.
di Guan, C., Li, P., Riggs, P. D., and Inouye, H. (1988) Vectors that facilitate the expression and purification of foreign peptides in Escherichia coli by fusion to maltose-binding protein. Gene 67, 21–30.
LaVallie, E. R., DiBlasio, E. A., Kovacic, S., Grant, K. L., Schendel, P. F., and McCoy, J. M. (1993) A thioredoxin gene fusion expression system that circumvents inclusion body formation in the E. coli cytoplasm. Bio/Technology 11, 187–193.
Davis, G. D., Elisee, C., Newham, D. M., and Harrison, R. G. (1999) New fusion protein systems designed to give soluble expression in Escherichia coli. Biotech. Bioeng. 65, 382–388.
Wilkinson, D. L. and Harrison, R. G. (1991) Predicting the solubility of recombinant proteins in Escherichia coli. Bio/Technology 9, 443–448.
Harrison, R. G. (2000) Expression of soluble heterologous proteins via fusion with NusA protein. inNovations (newsletter of Novagen, Inc., Madison, WI), June, 4–7.
Davis, G. D. and Harrison, R. G. (1998) Rapid screening of fusion protein recombinants by measuring effects of protein overexpression on cell growth. BioTechniques 24, 360–362.
Hofmann, M. A. and Brian, D. A. (1991) Sequencing PCR DNA amplified directly from a bacterial colony. Biotechniques 11, 30–31.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2003 Humana Press Inc.
About this protocol
Cite this protocol
Davis, G.D., Harrison, R.G. (2003). Discovery of New Fusion Protein Systems Designed to Enhance Solubility in E. coli . In: Vaillancourt, P.E. (eds) E. coliGene Expression Protocols. Methods in Molecular Biology™, vol 205. Humana Press. https://doi.org/10.1385/1-59259-301-1:141
Download citation
DOI: https://doi.org/10.1385/1-59259-301-1:141
Publisher Name: Humana Press
Print ISBN: 978-1-58829-008-3
Online ISBN: 978-1-59259-301-9
eBook Packages: Springer Protocols