ArticleActa crystallographica. Section F, Structural biology and crystallization communications2011
Expression of proteins in Escherichia coli as fusions with maltose-binding protein to rescue non-expressed targets in a high-throughput protein-expression and purification pipeline.
Article in Acta crystallographica. Section F, Structural biology and crystallization communications, 2011. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 23 papers.
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23 citing papers in PubMed, 29 citations in OpenAlex.
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- TIR domains of TLR family-from the cell culture to the protein sample for structural studies.PloS one · 2024Article
- Structural analysis and molecular substrate recognition properties ofFrontiers in plant science · 2023Article
- Enhanced serodiagnosis of melioidosis by indirect ELISA using the chimeric protein rGroEL-FLAG300 as an antigen.BMC infectious diseases · 2022Article
- FN3-based monobodies selective for the receptor binding domain of the SARS-CoV-2 spike protein.New biotechnology · 2021Article
- Directed Evolution of a Cp*RhChembiochem : a European journal of chemical biology · 2021Article
- Improvement of Soluble Production of Reteplase inAdvanced biomedical research · 2019Article
- Fusion proteins towards fungi and bacteria in plant protection.Microbiology (Reading, England) · 2018Article
- DNA-binding properties of FOXP3 transcription factor.Acta biochimica et biophysica Sinica · 2017Article
- Identification and Validation of Small-Gatekeeper Kinases as Drug Targets in Giardia lamblia.PLoS neglected tropical diseases · 2016Article
- High-throughput recombinant protein expression in Escherichia coli: current status and future perspectives.Open biology · 2016Review
- A dual protease approach for expression and affinity purification of recombinant proteins.Analytical biochemistry · 2016Article
- From Genome to Structure and Back Again: A Family Portrait of the Transcarbamylases.International journal of molecular sciences · 2015Review
- Positional effects of fusion partners on the yield and solubility of MBP fusion proteins.Protein expression and purification · 2015Article
- Seneca Valley Virus 3Cpro Substrate Optimization Yields Efficient Substrates for Use in Peptide-Prodrug Therapy.PloS one · 2015Article
- Article
- Several affinity tags commonly used in chromatographic purification.Journal of analytical methods in chemistry · 2013Review
- Microarray-based method for screening of immunogenic proteins from bacteria.Journal of nanobiotechnology · 2012Article
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Authors and funding
6 authors at 1 institution in 1 country.
Funding
Abstract
Despite recent advances, the expression of heterologous proteins in Escherichia coli for crystallization remains a nontrivial challenge. The present study investigates the efficacy of maltose-binding protein (MBP) fusion as a general strategy for rescuing the expression of target proteins. From a group of sequence-verified clones with undetectable levels of protein expression in an E. coli T7 expression system, 95 clones representing 16 phylogenetically diverse organisms were selected for recloning into a chimeric expression vector with an N-terminal histidine-tagged MBP. PCR-amplified inserts were annealed into an identical ligation-independent cloning region in an MBP-fusion vector and were analyzed for expression and solubility by high-throughput nickel-affinity binding. This approach yielded detectable expression of 72% of the clones; soluble expression was visible in 62%. However, the solubility of most proteins was marginal to poor upon cleavage of the MBP tag. This study offers large-scale evidence that MBP can improve the soluble expression of previously non-expressing proteins from a variety of eukaryotic and prokaryotic organisms. While the behavior of the cleaved proteins was disappointing, further refinements in MBP tagging may permit the more widespread use of MBP-fusion proteins in crystallographic studies.
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