ArticleProtein science : a publication of the Protein Society2009
Genetic selection system for improving recombinant membrane protein expression in E. coli.
Article in Protein science : a publication of the Protein Society, 2009. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 30 papers.
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Who cites it
30 citing papers in PubMed, 61 citations in OpenAlex.
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- Directed Evolution of Soluble α-1,2-Fucosyltransferase Using Kanamycin Resistance Protein as a Phenotypic Reporter for Efficient Production of 2'-Fucosyllactose.Journal of microbiology and biotechnology · 2022Article
- Smoothing membrane protein structure determination by initial upstream stage improvements.Applied microbiology and biotechnology · 2019Review
- Functionality of membrane proteins overexpressed and purified from E. coli is highly dependent upon the strain.Scientific reports · 2019Article
- Mutations in sigma 70 transcription factor improves expression of functional eukaryotic membrane proteins in Escherichia coli.Scientific reports · 2019Article
- Expression of Leptospira membrane proteins Signal Peptidase (SP) and Leptospira Endostatin like A (Len A) in BL-21(DE3) is toxic to the host cells.Journal, genetic engineering & biotechnology · 2018Article
- Heterologous Expression, Purification and Characterization of an Oligopeptidase A from the Pathogen Leptospira interrogans.Molecular biotechnology · 2018Article
- Improving membrane protein expression and function using genomic edits.Scientific reports · 2017Article
- Isolation and characterization of the E. coli membrane protein production strain Mutant56(DE3).Scientific reports · 2017Article
- Isolating Escherichia coli strains for recombinant protein production.Cellular and molecular life sciences : CMLS · 2017Review
- Production of FAME biodiesel in E. coli by direct methylation with an insect enzyme.Scientific reports · 2016Article
- Comparative genomics and experimental evolution of Escherichia coli BL21(DE3) strains reveal the landscape of toxicity escape from membrane protein overproduction.Scientific reports · 2015Article
- Dual-topology insertion of a dual-topology membrane protein.Nature communications · 2015Article
- A synthetic biochemistry system for the in vitro production of isoprene from glycolysis intermediates.Protein science : a publication of the Protein Society · 2014Article
- Membrane proteins can have high kinetic stability.Journal of the American Chemical Society · 2013Article
- Multi-copy genes that enhance the yield of mammalian G protein-coupled receptors in Escherichia coli.Metabolic engineering · 2012Article
- Combinatorial approaches for inverse metabolic engineering applications.Computational and structural biotechnology journal · 2012Review
- Physiological response to membrane protein overexpression in E. coli.Molecular & cellular proteomics : MCP · 2011Article
- A high-throughput immobilized bead screen for stable proteins and multi-protein complexes.Protein engineering, design & selection : PEDS · 2011Article
- Overexpression and purification of U24 from human herpesvirus type-6 in E. coli: unconventional use of oxidizing environments with a maltose binding protein-hexahistine dual tag to enhance membrane protein yield.Microbial cell factories · 2011Article
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Authors and funding
8 authors at 3 institutions in 1 country.
Funding
Abstract
A major barrier to the physical characterization and structure determination of membrane proteins is low yield in recombinant expression. To address this problem, we have designed a selection strategy to isolate mutant strains of Escherichia coli that improve the expression of a targeted membrane protein. In this method, the coding sequence of the membrane protein of interest is fused to a C-terminal selectable marker, so that the production of the selectable marker and survival on selective media is linked to expression of the targeted membrane protein. Thus, mutant strains with improved expression properties can be directly selected. We also introduce a rapid method for curing isolated strains of the plasmids used during the selection process, in which the plasmids are removed by in vivo digestion with the homing endonuclease I-CreI. We tested this selection system on a rhomboid family protein from Mycobacterium tuberculosis (Rv1337) and were able to isolate mutants, which we call EXP strains, with up to 75-fold increased expression. The EXP strains also improve the expression of other membrane proteins that were not the target of selection, in one case roughly 90-fold.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.