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usp2 catalytic domain expression construct  (Addgene inc)


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    Addgene inc usp2 catalytic domain expression construct
    Fluorescence polarization-based IsoMim assay principle and optimization. A , crystal structure of the catalytic core domain of USP11, depicted as a grey surface representation, in complex with Ub-GGG (in purple cartoon representation with the GGG extension colored in black ; PDB code: 8OYP ) B , schematic depiction of the probe design and assay principle. Representation of the DiUb 3G -FM substrate with the predicted Ub L73P -Ub structure (Alphafold ) in cartoon representation shown in purple and the C-terminal GGGC extension labeled with Fluorescein-5-maleimide (FM) shown as a chemical structure. The C-terminal ubiquitin moiety, the P1 position according to canonical protease nomenclature, is boxed and shaded in purple and forms the basis of a Ub 3G -FM probe. The GGGC extension is highlighted by a black-rimmed box . Upon DUB cleavage of DiUb 3G -FM, the rotational freedom of the liberated GGGC-FM moiety increases, leading to depolarization of the incident polarized light and a decrease in FP signal. C , SDS-PAGE gel analysis comparing mono-ubiquitin with the generated substrate DiUb 3G -FM, in the absence and presence of 50 nM <t>USP2</t> after 30 min incubation, and unlabeled DiUb 3G . D , comparison of the FP signal from DiUb 3G -FM (10 nM) with free fluorescein (10 nM) demonstrating the assay window. E , converted curves for 10 nM USP2, USP2 C276A and heat inactivated USP2 (USP ia ) showing the FP change. F , representative progress curves for USP2, USP2 C276A and heat inactivated USP2 (USP ia ) in comparison with the probe alone control ( top inset ). Rates of reaction at three different USP2 concentrations with 10 nM DiUb 3G -FM ( bottom ). Data points represent the mean ± SD of three independent experiments.
    Usp2 Catalytic Domain Expression Construct, supplied by Addgene inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/usp2 catalytic domain expression construct/product/Addgene inc
    Average 90 stars, based on 1 article reviews
    usp2 catalytic domain expression construct - by Bioz Stars, 2026-05
    90/100 stars

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    1) Product Images from "A versatile fluorescence polarization-based deubiquitination assay using an isopeptide bond substrate mimetic (IsoMim)"

    Article Title: A versatile fluorescence polarization-based deubiquitination assay using an isopeptide bond substrate mimetic (IsoMim)

    Journal: The Journal of Biological Chemistry

    doi: 10.1016/j.jbc.2025.110342

    Fluorescence polarization-based IsoMim assay principle and optimization. A , crystal structure of the catalytic core domain of USP11, depicted as a grey surface representation, in complex with Ub-GGG (in purple cartoon representation with the GGG extension colored in black ; PDB code: 8OYP ) B , schematic depiction of the probe design and assay principle. Representation of the DiUb 3G -FM substrate with the predicted Ub L73P -Ub structure (Alphafold ) in cartoon representation shown in purple and the C-terminal GGGC extension labeled with Fluorescein-5-maleimide (FM) shown as a chemical structure. The C-terminal ubiquitin moiety, the P1 position according to canonical protease nomenclature, is boxed and shaded in purple and forms the basis of a Ub 3G -FM probe. The GGGC extension is highlighted by a black-rimmed box . Upon DUB cleavage of DiUb 3G -FM, the rotational freedom of the liberated GGGC-FM moiety increases, leading to depolarization of the incident polarized light and a decrease in FP signal. C , SDS-PAGE gel analysis comparing mono-ubiquitin with the generated substrate DiUb 3G -FM, in the absence and presence of 50 nM USP2 after 30 min incubation, and unlabeled DiUb 3G . D , comparison of the FP signal from DiUb 3G -FM (10 nM) with free fluorescein (10 nM) demonstrating the assay window. E , converted curves for 10 nM USP2, USP2 C276A and heat inactivated USP2 (USP ia ) showing the FP change. F , representative progress curves for USP2, USP2 C276A and heat inactivated USP2 (USP ia ) in comparison with the probe alone control ( top inset ). Rates of reaction at three different USP2 concentrations with 10 nM DiUb 3G -FM ( bottom ). Data points represent the mean ± SD of three independent experiments.
    Figure Legend Snippet: Fluorescence polarization-based IsoMim assay principle and optimization. A , crystal structure of the catalytic core domain of USP11, depicted as a grey surface representation, in complex with Ub-GGG (in purple cartoon representation with the GGG extension colored in black ; PDB code: 8OYP ) B , schematic depiction of the probe design and assay principle. Representation of the DiUb 3G -FM substrate with the predicted Ub L73P -Ub structure (Alphafold ) in cartoon representation shown in purple and the C-terminal GGGC extension labeled with Fluorescein-5-maleimide (FM) shown as a chemical structure. The C-terminal ubiquitin moiety, the P1 position according to canonical protease nomenclature, is boxed and shaded in purple and forms the basis of a Ub 3G -FM probe. The GGGC extension is highlighted by a black-rimmed box . Upon DUB cleavage of DiUb 3G -FM, the rotational freedom of the liberated GGGC-FM moiety increases, leading to depolarization of the incident polarized light and a decrease in FP signal. C , SDS-PAGE gel analysis comparing mono-ubiquitin with the generated substrate DiUb 3G -FM, in the absence and presence of 50 nM USP2 after 30 min incubation, and unlabeled DiUb 3G . D , comparison of the FP signal from DiUb 3G -FM (10 nM) with free fluorescein (10 nM) demonstrating the assay window. E , converted curves for 10 nM USP2, USP2 C276A and heat inactivated USP2 (USP ia ) showing the FP change. F , representative progress curves for USP2, USP2 C276A and heat inactivated USP2 (USP ia ) in comparison with the probe alone control ( top inset ). Rates of reaction at three different USP2 concentrations with 10 nM DiUb 3G -FM ( bottom ). Data points represent the mean ± SD of three independent experiments.

    Techniques Used: Fluorescence, Labeling, Ubiquitin Proteomics, SDS Page, Generated, Incubation, Comparison, Control

    Dose response curves of different DUBs and PR-619 as the inhibitor. A , dose–response curves for inhibition by pan-inhibitor PR-619 for the catalytic core domains of USP4, USP15, USP2 and UCHL3. Data were fitted using non-linear regression in GraphPad Prism, and data points show mean ± SD from three independent experiments. B , pIC50 values of PR-619 DUB inhibition (error = SE; n = 3). C , proof-of principle high-throughput screening results using USP4-D1D2 and PR-619 in a 384-well plate layout. The DiUb 3G -FM reagent alone and selected wells together with USP4-D1D2 served as control groups, measured as quadruplicates on the plate as indicated in . PR-619 was added in random places in quadruplicate (plate layout displayed in ). Highlighted in light green are the data from wells where USP4-D1D2 was incubated with PR-619. Dashed lines indicate 0% and 50% of inhibition.
    Figure Legend Snippet: Dose response curves of different DUBs and PR-619 as the inhibitor. A , dose–response curves for inhibition by pan-inhibitor PR-619 for the catalytic core domains of USP4, USP15, USP2 and UCHL3. Data were fitted using non-linear regression in GraphPad Prism, and data points show mean ± SD from three independent experiments. B , pIC50 values of PR-619 DUB inhibition (error = SE; n = 3). C , proof-of principle high-throughput screening results using USP4-D1D2 and PR-619 in a 384-well plate layout. The DiUb 3G -FM reagent alone and selected wells together with USP4-D1D2 served as control groups, measured as quadruplicates on the plate as indicated in . PR-619 was added in random places in quadruplicate (plate layout displayed in ). Highlighted in light green are the data from wells where USP4-D1D2 was incubated with PR-619. Dashed lines indicate 0% and 50% of inhibition.

    Techniques Used: Inhibition, High Throughput Screening Assay, Control, Incubation



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    usp2 catalytic domain expression construct  (Addgene inc)
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    Addgene inc usp2 catalytic domain expression construct
    Fluorescence polarization-based IsoMim assay principle and optimization. A , crystal structure of the catalytic core domain of USP11, depicted as a grey surface representation, in complex with Ub-GGG (in purple cartoon representation with the GGG extension colored in black ; PDB code: 8OYP ) B , schematic depiction of the probe design and assay principle. Representation of the DiUb 3G -FM substrate with the predicted Ub L73P -Ub structure (Alphafold ) in cartoon representation shown in purple and the C-terminal GGGC extension labeled with Fluorescein-5-maleimide (FM) shown as a chemical structure. The C-terminal ubiquitin moiety, the P1 position according to canonical protease nomenclature, is boxed and shaded in purple and forms the basis of a Ub 3G -FM probe. The GGGC extension is highlighted by a black-rimmed box . Upon DUB cleavage of DiUb 3G -FM, the rotational freedom of the liberated GGGC-FM moiety increases, leading to depolarization of the incident polarized light and a decrease in FP signal. C , SDS-PAGE gel analysis comparing mono-ubiquitin with the generated substrate DiUb 3G -FM, in the absence and presence of 50 nM <t>USP2</t> after 30 min incubation, and unlabeled DiUb 3G . D , comparison of the FP signal from DiUb 3G -FM (10 nM) with free fluorescein (10 nM) demonstrating the assay window. E , converted curves for 10 nM USP2, USP2 C276A and heat inactivated USP2 (USP ia ) showing the FP change. F , representative progress curves for USP2, USP2 C276A and heat inactivated USP2 (USP ia ) in comparison with the probe alone control ( top inset ). Rates of reaction at three different USP2 concentrations with 10 nM DiUb 3G -FM ( bottom ). Data points represent the mean ± SD of three independent experiments.
    Usp2 Catalytic Domain Expression Construct, supplied by Addgene inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/usp2 catalytic domain expression construct/product/Addgene inc
    Average 90 stars, based on 1 article reviews
    usp2 catalytic domain expression construct - by Bioz Stars, 2026-05
    90/100 stars
    		  Buy from Supplier

    Image Search Results


    Fluorescence polarization-based IsoMim assay principle and optimization. A , crystal structure of the catalytic core domain of USP11, depicted as a grey surface representation, in complex with Ub-GGG (in purple cartoon representation with the GGG extension colored in black ; PDB code: 8OYP ) B , schematic depiction of the probe design and assay principle. Representation of the DiUb 3G -FM substrate with the predicted Ub L73P -Ub structure (Alphafold ) in cartoon representation shown in purple and the C-terminal GGGC extension labeled with Fluorescein-5-maleimide (FM) shown as a chemical structure. The C-terminal ubiquitin moiety, the P1 position according to canonical protease nomenclature, is boxed and shaded in purple and forms the basis of a Ub 3G -FM probe. The GGGC extension is highlighted by a black-rimmed box . Upon DUB cleavage of DiUb 3G -FM, the rotational freedom of the liberated GGGC-FM moiety increases, leading to depolarization of the incident polarized light and a decrease in FP signal. C , SDS-PAGE gel analysis comparing mono-ubiquitin with the generated substrate DiUb 3G -FM, in the absence and presence of 50 nM USP2 after 30 min incubation, and unlabeled DiUb 3G . D , comparison of the FP signal from DiUb 3G -FM (10 nM) with free fluorescein (10 nM) demonstrating the assay window. E , converted curves for 10 nM USP2, USP2 C276A and heat inactivated USP2 (USP ia ) showing the FP change. F , representative progress curves for USP2, USP2 C276A and heat inactivated USP2 (USP ia ) in comparison with the probe alone control ( top inset ). Rates of reaction at three different USP2 concentrations with 10 nM DiUb 3G -FM ( bottom ). Data points represent the mean ± SD of three independent experiments.

    Journal: The Journal of Biological Chemistry

    Article Title: A versatile fluorescence polarization-based deubiquitination assay using an isopeptide bond substrate mimetic (IsoMim)

    doi: 10.1016/j.jbc.2025.110342

    Figure Lengend Snippet: Fluorescence polarization-based IsoMim assay principle and optimization. A , crystal structure of the catalytic core domain of USP11, depicted as a grey surface representation, in complex with Ub-GGG (in purple cartoon representation with the GGG extension colored in black ; PDB code: 8OYP ) B , schematic depiction of the probe design and assay principle. Representation of the DiUb 3G -FM substrate with the predicted Ub L73P -Ub structure (Alphafold ) in cartoon representation shown in purple and the C-terminal GGGC extension labeled with Fluorescein-5-maleimide (FM) shown as a chemical structure. The C-terminal ubiquitin moiety, the P1 position according to canonical protease nomenclature, is boxed and shaded in purple and forms the basis of a Ub 3G -FM probe. The GGGC extension is highlighted by a black-rimmed box . Upon DUB cleavage of DiUb 3G -FM, the rotational freedom of the liberated GGGC-FM moiety increases, leading to depolarization of the incident polarized light and a decrease in FP signal. C , SDS-PAGE gel analysis comparing mono-ubiquitin with the generated substrate DiUb 3G -FM, in the absence and presence of 50 nM USP2 after 30 min incubation, and unlabeled DiUb 3G . D , comparison of the FP signal from DiUb 3G -FM (10 nM) with free fluorescein (10 nM) demonstrating the assay window. E , converted curves for 10 nM USP2, USP2 C276A and heat inactivated USP2 (USP ia ) showing the FP change. F , representative progress curves for USP2, USP2 C276A and heat inactivated USP2 (USP ia ) in comparison with the probe alone control ( top inset ). Rates of reaction at three different USP2 concentrations with 10 nM DiUb 3G -FM ( bottom ). Data points represent the mean ± SD of three independent experiments.

    Article Snippet: The USP2 catalytic domain expression construct as used for the structure determination of a covalent Ubiquitin-USP2 complex (PDB code: 2IBI ) was a gift from Cheryl Arrowsmith (Addgene plasmid # 36894; http://n2t.net/addgene:36894 ; RRID: Addgene 36,894).

    Techniques: Fluorescence, Labeling, Ubiquitin Proteomics, SDS Page, Generated, Incubation, Comparison, Control

    Dose response curves of different DUBs and PR-619 as the inhibitor. A , dose–response curves for inhibition by pan-inhibitor PR-619 for the catalytic core domains of USP4, USP15, USP2 and UCHL3. Data were fitted using non-linear regression in GraphPad Prism, and data points show mean ± SD from three independent experiments. B , pIC50 values of PR-619 DUB inhibition (error = SE; n = 3). C , proof-of principle high-throughput screening results using USP4-D1D2 and PR-619 in a 384-well plate layout. The DiUb 3G -FM reagent alone and selected wells together with USP4-D1D2 served as control groups, measured as quadruplicates on the plate as indicated in . PR-619 was added in random places in quadruplicate (plate layout displayed in ). Highlighted in light green are the data from wells where USP4-D1D2 was incubated with PR-619. Dashed lines indicate 0% and 50% of inhibition.

    Journal: The Journal of Biological Chemistry

    Article Title: A versatile fluorescence polarization-based deubiquitination assay using an isopeptide bond substrate mimetic (IsoMim)

    doi: 10.1016/j.jbc.2025.110342

    Figure Lengend Snippet: Dose response curves of different DUBs and PR-619 as the inhibitor. A , dose–response curves for inhibition by pan-inhibitor PR-619 for the catalytic core domains of USP4, USP15, USP2 and UCHL3. Data were fitted using non-linear regression in GraphPad Prism, and data points show mean ± SD from three independent experiments. B , pIC50 values of PR-619 DUB inhibition (error = SE; n = 3). C , proof-of principle high-throughput screening results using USP4-D1D2 and PR-619 in a 384-well plate layout. The DiUb 3G -FM reagent alone and selected wells together with USP4-D1D2 served as control groups, measured as quadruplicates on the plate as indicated in . PR-619 was added in random places in quadruplicate (plate layout displayed in ). Highlighted in light green are the data from wells where USP4-D1D2 was incubated with PR-619. Dashed lines indicate 0% and 50% of inhibition.

    Article Snippet: The USP2 catalytic domain expression construct as used for the structure determination of a covalent Ubiquitin-USP2 complex (PDB code: 2IBI ) was a gift from Cheryl Arrowsmith (Addgene plasmid # 36894; http://n2t.net/addgene:36894 ; RRID: Addgene 36,894).

    Techniques: Inhibition, High Throughput Screening Assay, Control, Incubation