ArticleEukaryotic cell2012
Plasmid vectors for proteomic analyses in Giardia: purification of virulence factors and analysis of the proteasome.
Article in Eukaryotic cell, 2012. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 20 papers.
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Who cites it
20 citing papers in PubMed, 41 citations in OpenAlex.
- Investigating Antiprotozoal Chemotherapies with Novel Proteomic Tools-Chances and Limitations: A Critical Review.International journal of molecular sciences · 2024Review
- Harnessing the power of new genetic tools to illuminate Giardia biology and pathogenesis.Genetics · 2024Review
- A myeloid leukemia factor homolog is involved in tolerance to stresses and stress-induced protein metabolism in Giardia lamblia.Biology direct · 2023Article
- A Detailed Gene Expression Map ofGenes · 2021Article
- Article
- Methionyl-tRNA synthetase inhibitor has potent in vivo activity in a novel Giardia lamblia luciferase murine infection model.The Journal of antimicrobial chemotherapy · 2020Article
- High Cysteine Membrane Proteins (HCMPs) Are Up-Regulated DuringFrontiers in genetics · 2020Article
- Recent advances in functional research in Giardia intestinalis.Advances in parasitology · 2020Review
- Article
- Secreted Giardia intestinalis cysteine proteases disrupt intestinal epithelial cell junctional complexes and degrade chemokines.Virulence · 2018Article
- Coordinated Changes in Gene Expression Throughout Encystation of Giardia intestinalis.PLoS neglected tropical diseases · 2016Article
- Comparative genomic analyses of freshly isolated Giardia intestinalis assemblage A isolates.BMC genomics · 2015Article
- A reduced VWA domain-containing proteasomal ubiquitin receptor of Giardia lamblia localizes to the flagellar pore regions in microtubule-dependent manner.Parasites & vectors · 2015Article
- Identification of obscure yet conserved actin-associated proteins in Giardia lamblia.Eukaryotic cell · 2014Article
- The genome of Spironucleus salmonicida highlights a fish pathogen adapted to fluctuating environments.PLoS genetics · 2014Article
- The role of arginine and arginine-metabolizing enzymes during Giardia - host cell interactions in vitro.BMC microbiology · 2013Article
- Hydrogenosomes in the diplomonad Spironucleus salmonicida.Nature communications · 2013Article
- Stable transfection of the diplomonad parasite Spironucleus salmonicida.Eukaryotic cell · 2012Article
- Arginine consumption by the intestinal parasite Giardia intestinalis reduces proliferation of intestinal epithelial cells.PloS one · 2012Article
- Article
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Authors and funding
5 authors at 2 institutions in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
In recent years, proteomics has come of age with the development of efficient tools for purification, identification, and characterization of gene products predicted by genome projects. The intestinal protozoan Giardia intestinalis can be transfected, but there is only a limited set of vectors available, and most of them are not user friendly. This work delineates the construction of a suite of cassette-based expression vectors for use in Giardia. Expression is provided by the strong constitutive ornithine carbamoyltransferase (OCT) promoter, and tagging is possible in both N- and C-terminal configurations. Taken together, the vectors are capable of providing protein localization and production of recombinant proteins, followed by efficient purification by a novel affinity tag combination, streptavidin binding peptide-glutathione S-transferase (SBP-GST). The option of removing the tags from purified proteins was provided by the inclusion of a PreScission protease site. The efficiency and feasibility of producing and purifying endogenous recombinant Giardia proteins with the developed vectors was demonstrated by the purification of active recombinant arginine deiminase (ADI) and OCT from stably transfected trophozoites. Moreover, we describe the tagging, purification by StrepTactin affinity chromatography, and compositional analysis by mass spectrometry of the G. intestinalis 26S proteasome by employing the Strep II-FLAG-tandem affinity purification (SF-TAP) tag. This is the first report of efficient production and purification of recombinant proteins in and from Giardia, which will allow the study of specific parasite proteins and protein complexes.
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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.