Pick a Tag and Explore the Functions of Your Pet Protein

Vandemoortele, Giel; Eyckerman, Sven; Gevaert, Kris

doi:10.1016/j.tibtech.2019.03.016

Cited by 54 publications

(45 citation statements)

References 87 publications

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“…In the last 5 years, there have been many reports describing new fusion partners and affinity tags, including in‐depth reviews . In the area of small peptide tags, Ojima‐Kato et al reported that the addition of the sequence coding for serine‐lysine‐isoleucine‐lysine (SKIK tag) after the initial N‐terminal methionine codon markedly improves the expression of recombinant proteins .…”

Section: Advances In Plasmid Designmentioning

confidence: 99%

New tools for recombinant protein production in Escherichia coli: A 5‐year update

2019

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The production of proteins in sufficient amounts is key for their study or use as biotherapeutic agents. Escherichia coli is the host of choice for recombinant protein production given its fast growth, easy manipulation, and cost-effectiveness. As such, its protein production capabilities are continuously being improved. Also, the associated tools (such as plasmids and cultivation conditions) are subject of ongoing research to optimize product yield. In this work, we review the latest advances in recombinant protein production in E. coli.

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Section: Advances In Plasmid Designmentioning

confidence: 99%

New tools for recombinant protein production in Escherichia coli: A 5‐year update

2019

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“…To overcome these limitations and have in hand a reliable tool for the study of RanBP9 in physiological and pathological paradigms in vivo, we decided to use CRISPR/Cas9 and generate RanBP9-TT mice by the addition of a double V5-HA tag at the C-terminus of RanBP9. Biochemical tags are increasingly an excellent solution for detection problems and are widely employed for research purposes 19 .…”

Section: Discussionmentioning

confidence: 99%

Tagging enhances histochemical and biochemical detection of Ran Binding Protein 9 in vivo and reveals its interaction with Nucleolin

Soliman

Stark

Gardner

et al. 2020

Sci Rep

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The lack of tools to reliably detect RanBP9 in vivo has significantly hampered progress in understanding the biological functions of this scaffold protein. We report here the generation of a novel mouse strain, RanBP9-TT, in which the endogenous protein is fused with a double (V5-HA) epitope tag at the C-terminus. We show that the double tag does not interfere with the essential functions of RanBP9. In contrast to RanBP9 constitutive knock-out animals, RanBP9-TT mice are viable, fertile and do not show any obvious phenotype. The V5-HA tag allows unequivocal detection of RanBP9 both by IHC and WB. Importantly, immunoprecipitation and mass spectrometry analyses reveal that the tagged protein pulls down known interactors of wild type RanBP9. Thanks to the increased detection power, we are also unveiling a previously unknown interaction with Nucleolin, a protein proposed as an ideal target for cancer treatment. In summary, we report the generation of a new mouse line in which RanBP9 expression and interactions can be reliably studied by the use of commercially available αtag antibodies. The use of this line will help to overcome some of the existing limitations in the study of RanBP9 and potentially unveil unknown functions of this protein in vivo such as those linked to Nucleolin.

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“…To overcome these limitations and have in hand a reliable in vivo tool for the study of RanBP9 in physiological and pathological paradigms, we decided to use CRISPR/Cas9 and generate RanBP9-TT mice by the addition of a double V5-HA tag at the C-terminus of RanBP9. Increasingly, biochemical tags are an excellent solution for detection problems and are widely employed for research purposes 19 .…”

Section: Discussionmentioning

confidence: 99%

Tagging enhances histochemical and biochemical detection of Ran Binding Protein 9 in vivo and reveals its interaction with Nucleolin

Soliman

Stark

Gardner

et al. 2019

Preprint

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AbstractThe lack of tools to reliably detect RanBP9 in vivo has significantly hampered progress in understanding the biological functions of this scaffold protein. We report here the generation of a novel mouse strain, RanBP9-TT, in which the endogenous protein is fused with a double (V5-HA) epitope tag at the C-terminus. We show that the double tag does not interfere with the essential functions of RanBP9. In contrast to RanBP9 constitutive knock-out animals, RanBP9-TT mice are viable, fertile and do not show any obvious phenotype. The V5-HA tag allows unequivocal detection of RanBP9 both by IHC and WB. Importantly, immunoprecipitation and mass spectrometry analyses reveal that the tagged protein pulls down known interactors of wild type RanBP9. Thanks to the increased detection power, we are also unveiling a novel interaction with Nucleolin, a prominent nucleolar protein.In summary, we report the generation of a new mouse line in which RanBP9 expression and interactions can be reliably studied by the use of commercially available αtag antibodies. The use of this line will help to overcome some of the existing limitations in the study of RanBP9 and potentially reveal novel functions of this protein in vivo such as those linked to Nucleolin.

show abstract

Pick a Tag and Explore the Functions of Your Pet Protein

Cited by 54 publications

References 87 publications

New tools for recombinant protein production in Escherichia coli: A 5‐year update

New tools for recombinant protein production in Escherichia coli: A 5‐year update

Tagging enhances histochemical and biochemical detection of Ran Binding Protein 9 in vivo and reveals its interaction with Nucleolin

Tagging enhances histochemical and biochemical detection of Ran Binding Protein 9 in vivo and reveals its interaction with Nucleolin

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