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e2 ubch5b (e2; boston biochem)  (Boston Biochem)


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    Structured Review

    Boston Biochem e2 ubch5b (e2; boston biochem)
    E2 Ubch5b (E2; Boston Biochem), supplied by Boston Biochem, 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/product/e2+ubch5b+(e2%3B+boston+biochem)/ubch5b+enzyme/pmc11101318-220-27-30
    Average 90 stars, based on 1 article reviews
    e2 ubch5b (e2; boston biochem) - by Bioz Stars, 2026-09
    90/100 stars

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    Ubiquitin Proteomics:

    Article Title: Post-translational regulation of the Numb/Notch pathway in neurogenesis and cancer by Dlk2
    Article Snippet: .. Reactions contained Tris 40 mM pH 7.4, 5mM Mg-ATP solution (Boston Biochem, B-20), 0.5 mM DTT, 0.6 μM ubiquitin (Boston Biochem, U-120), 225 nM E1-UBE-1 (Boston Biochem, E-305), 1.25 μM E2-UbcH5b (Boston Biochem, E3-622), and 2.5 μM p53 (Boston Biochem, SP-450). .. Reactions were started using E3 ligase MDM2/HDM2 400 ng (Boston Biochem, E3-204), incubated for 3 hours at 30°C in a water bath.

    Article Title: Suppression of ITPKB degradation by Trim25 confers TMZ resistance in glioblastoma through ROS homeostasis
    Article Snippet: The E3 ligase V5-Trim25 was mixed with 10 U of FastAPTM Thermosensitive alkaline phosphatase (Thermo Scientific, #EF0654). .. The E3 ligase complex was then mixed with 2 μg Flag-ITPKB purified from HEK293T cells and reacted with 15 μg wild-type ubiquitin (Boston Biochem, U-100H-10M), 550 ng of E1 (Uba1, Boston Biochem, E-305-025), and 850 ng of E2 (UbcH5b, Boston Biochem, E2-622-100). ..

    Article Title: Deubiquitination of BES1 by UBP12/UBP13 pro- motes brassinosteroid signaling and plant growth
    Article Snippet: .. Plant Commun Ubiquitination and deubiquitination assays in vitro To obtain polyubiquitinated MBP-BES1-myc in vitro, 500 ng of MBPBES1-myc was mixed with 55 nM of human UBE1 (E1; Boston Biochem, cat. no. E-304), 166 nM of human UbcH5b (E2; Boston Biochem, cat. no. E2-622), 500 ng of GST-SINAT2 (E3), and 2 mg of histidine-tagged ubiquitin (His-ubiquitin, 10.7 kDa) in 30 ml of a ubiquitination buffer (50 mM Tris [pH 7.4], 5 mM MgCl2, 2 mM DTT, and 2 mM ATP). ..

    other:

    Article Title: Specificities and redundancies in the NEL family of bacterial E3 ubiquitin ligases of Salmonella enterica serovar Typhimurium
    Article Snippet: Ubiquitination reactions were carried out in a 20-μl mixture containing buffer A (25 mM Tris-HCl, pH 7.5, 50 mM NaCl, 5 mM ATP, 10 mM MgCl 2 , 0.1 mM DTT), 2 μg of HA-tagged ubiquitin, 0.25 μg of E1 (Boston Biochem, Cambridge, MA, USA) and 1 μg of E2 (human recombinant UbcH5b from Boston Biochem) in the presence or absence of ubiquitin ligases and substrates.

    Article Title: An alfalfa MYB-like transcriptional factor MsMYBH positively regulates alfalfa seedling drought resistance and undergoes MsWAV3-mediated degradation.
    Article Snippet: Briefly, In total, 100 ng UBA1 (E1; Boston Biochem), 200 ng of UbcH5b (E2; Boston Biochem), 10 ug of FLAG‐tagged ubiquitin (FLAG‐ Ub; Boston Biochem), 0.5 μg of GST‐MsMYBH, and 0.5 μg of MBP‐MsWAV3 were incubated in ubiquitination reaction buffer (50mM Tris‐HCl pH 7.5, 10mMMgCl2, 50mM ATP, and 20mM DTT) at 30°C for 2 h. After the reaction, the samples were added with 5 × protein loading buffer and then boiled at 95°C for 5min, which were subjected to immunoblotting with anti‐GST and anti‐MBP antibodies (Huaxingbio, 1:5,000 dilution).

    Article Title: Calmodulin-like protein MdCML15 interacts with MdBT2 to modulate iron homeostasis in apple
    Article Snippet: The reaction system (30 μl) comprised 200 ng HIS-MdbHLH104, 400 ng MdBT2–GFP or GFP, 500 ng MYC–MdCML15 or MYC, 25 ng UBE1 (E1; Boston Biochem), 50 ng E2 UbcH5b (E2; Boston Biochem), and 1 μg ubiquitin in a ubiquitination buffer (composed of 2 mM DTT, 50 mM Tris–HCl (pH 7.5), 5 mM MgCl 2 , and 2 mM ATP).

    Article Title: UV-B irradiation-activated E3 ligase GmILPA1 modulates gibberellin catabolism to increase plant height in soybean.
    Article Snippet: The other ingredients, including E1, E2 (UbcH5b/UBE2D2), and ubiquitin (Ub), were purchased from Boston Biochem (Cambridge, MA).

    Article Title: ArMYB89 and ArCOP1 interaction modulates anthocyanin biosynthesis in Acer rubrum leaves under low-temperature conditions.
    Article Snippet: Acer rubrum, a famous ornamental tree, produces bright red-coloured leaves because of the temperature decline from summer to autumn.. This process’s molecular mechanism is elusive, so we have investigated how anthocyanin biosynthesis is induced in A. rubrum leaves under low temperatures.. The results of low-temperature treatment under light and dark conditions showed that the low-temperature promoted anthocyanin accumulation in A. rubrum is light-dependent.

    Purification:

    Article Title: Suppression of ITPKB degradation by Trim25 confers TMZ resistance in glioblastoma through ROS homeostasis
    Article Snippet: The E3 ligase V5-Trim25 was mixed with 10 U of FastAPTM Thermosensitive alkaline phosphatase (Thermo Scientific, #EF0654). .. The E3 ligase complex was then mixed with 2 μg Flag-ITPKB purified from HEK293T cells and reacted with 15 μg wild-type ubiquitin (Boston Biochem, U-100H-10M), 550 ng of E1 (Uba1, Boston Biochem, E-305-025), and 850 ng of E2 (UbcH5b, Boston Biochem, E2-622-100). ..

    In Vitro:

    Article Title: Deubiquitination of BES1 by UBP12/UBP13 pro- motes brassinosteroid signaling and plant growth
    Article Snippet: .. Plant Commun Ubiquitination and deubiquitination assays in vitro To obtain polyubiquitinated MBP-BES1-myc in vitro, 500 ng of MBPBES1-myc was mixed with 55 nM of human UBE1 (E1; Boston Biochem, cat. no. E-304), 166 nM of human UbcH5b (E2; Boston Biochem, cat. no. E2-622), 500 ng of GST-SINAT2 (E3), and 2 mg of histidine-tagged ubiquitin (His-ubiquitin, 10.7 kDa) in 30 ml of a ubiquitination buffer (50 mM Tris [pH 7.4], 5 mM MgCl2, 2 mM DTT, and 2 mM ATP). ..



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    Phosphorylation of Trim25 at S100 is required for ITPKB ubiquitination. a Phosphorylation proteomics analysis of Trim25 in primary and recurrence GBM. b The phosphorylation level of Trim25 S100 site in GBM tissue. c Dephosphorylation of the E3 ligase Trim25 resulted in a more pronounced alleviation of ITPKB ubiquitination in a cell-free system compared to mock-treated Trim25. Dephosphorylated or mock-treated Trim25 was incubated with immunopurified Flag-ITPKB, ubiquitin, recombinant E1 (Uba1), and E2 (UbcH5b). ITPKB ubiquitination was then determined using a ubiquitination assay. d The interaction between ITPKB and Trim25 was affected by the Trim25 S100D phosphomimetic mutant and S100A dephosphorylation mutant. e The ubiquitination of ITPKB was altered when the S100 site was mutated to Asp (S100D) or Ala (S100A). HEK293T cells were co-transfected with the respective plasmids and treated with MG132 for 3 h after 48 h of transfection. Cell lysates were immunoprecipitated and blotted with specific antibodies for analysis. Statistical significance is shown as: * p < 0.01

    Journal: Signal Transduction and Targeted Therapy

    Article Title: Suppression of ITPKB degradation by Trim25 confers TMZ resistance in glioblastoma through ROS homeostasis

    doi: 10.1038/s41392-024-01763-x

    Figure Lengend Snippet: Phosphorylation of Trim25 at S100 is required for ITPKB ubiquitination. a Phosphorylation proteomics analysis of Trim25 in primary and recurrence GBM. b The phosphorylation level of Trim25 S100 site in GBM tissue. c Dephosphorylation of the E3 ligase Trim25 resulted in a more pronounced alleviation of ITPKB ubiquitination in a cell-free system compared to mock-treated Trim25. Dephosphorylated or mock-treated Trim25 was incubated with immunopurified Flag-ITPKB, ubiquitin, recombinant E1 (Uba1), and E2 (UbcH5b). ITPKB ubiquitination was then determined using a ubiquitination assay. d The interaction between ITPKB and Trim25 was affected by the Trim25 S100D phosphomimetic mutant and S100A dephosphorylation mutant. e The ubiquitination of ITPKB was altered when the S100 site was mutated to Asp (S100D) or Ala (S100A). HEK293T cells were co-transfected with the respective plasmids and treated with MG132 for 3 h after 48 h of transfection. Cell lysates were immunoprecipitated and blotted with specific antibodies for analysis. Statistical significance is shown as: * p < 0.01

    Article Snippet: The E3 ligase complex was then mixed with 2 μg Flag-ITPKB purified from HEK293T cells and reacted with 15 μg wild-type ubiquitin (Boston Biochem, U-100H-10M), 550 ng of E1 (Uba1, Boston Biochem, E-305-025), and 850 ng of E2 (UbcH5b, Boston Biochem, E2-622-100).

    Techniques: Phospho-proteomics, Ubiquitin Proteomics, De-Phosphorylation Assay, Incubation, Recombinant, Mutagenesis, Transfection, Immunoprecipitation