Expression screening of fusion partners from an E. coli genome for soluble expression of recombinant proteins in a cell-free protein synthesis system

While access to soluble recombinant proteins is essential for a number of proteome studies, preparation of purified functional proteins is often limited by the protein solubility. In this study, potent solubility-enhancing fusion partners were screened from the repertoire of endogenous E. coli prote...

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Published inPloS one Vol. 6; no. 11; p. e26875
Main Authors Ahn, Jin-Ho, Keum, Jung-Won, Kim, Dong-Myung
Format Journal Article
LanguageEnglish
Published United States Public Library of Science 02.11.2011
Public Library of Science (PLoS)
Subjects
Base Sequence
Biology
Cell-Free System
Chemical engineering
Chemistry
Cytokines
E coli
Efficiency
Escherichia coli
Escherichia coli - genetics
Freedom of speech
Gene expression
Genome, Bacterial
Genomes
Genomics
Kinases
Medical screening
Molecular Sequence Data
Open Reading Frames
Polymerase Chain Reaction
Polypeptides
Protein biosynthesis
Protein synthesis
Proteins
Proteomes
Recombinant proteins
Recombinant Proteins - genetics
Solubility
Transfer RNA
Yeast
United States > US
Massachusetts
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Summary:While access to soluble recombinant proteins is essential for a number of proteome studies, preparation of purified functional proteins is often limited by the protein solubility. In this study, potent solubility-enhancing fusion partners were screened from the repertoire of endogenous E. coli proteins. Based on the presumed correlation between the intracellular abundance and folding efficiency of proteins, PCR-amplified ORFs of a series of highly abundant E. coli proteins were fused with aggregation-prone heterologous proteins and then directly expressed for quantitative estimation of the expression efficiency of soluble translation products. Through two-step screening procedures involving the expression of 552 fusion constructs targeted against a series of cytokine proteins, we were able to discover a number of endogenous E. coli proteins that dramatically enhanced the soluble expression of the target proteins. This strategy of cell-free expression screening can be extended to quantitative, global analysis of genomic resources for various purposes.
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Current address: Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States of America
Current address: Department of Chemical Engineering, University of Massachusetts, Amherst, Massachusetts, United States of America
Conceived and designed the experiments: DK. Performed the experiments: JA JK. Analyzed the data: JA DK. Contributed reagents/materials/analysis tools: DK. Wrote the paper: JA DK.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0026875