Recombinant Human SIRT1, GST-tagged

• Cat.No.: SIRT1-462H
• Product Overview: Recombinant Human SIRT1 (amino acids 193- 741) was expressed in anE.coliexpression system with N-terminal GST tag. MW=87.2 kDa.

Specification

• Species: Human
• Source: E.coli
• Tag: GST
• Protein Length: 193-741 a.a.
• Description: Sirtuin 1 also known as NAD-dependent deacetylase sirtuin-1 is a protein that in humans is encoded by the SIRT1 gene. SIRT1 stands for sirtuin (silent mating type information regulation 2 homolog) 1 (S. cerevisiae), referring to the fact that its sirtuin homolog (biological equivalent across species) in yeast (S. cerevisiae) is Sir2. SIRT1 is an enzyme which deacetylates proteins that contribute to cellular regulation (reaction to stressors, longevity).
• Formulated In: 25 mM Tris-HCl, pH 8.0, 100 mM NaCl, 0.05% Tween-20 and 20% glycerol.
• Application: Useful for the study of enzyme kinetics, screening inhibitors, and selectivity profiling.
• Specific Activity: 8 pmol/min /μg. Assay condition: 25 mM Tris/Cl, pH8.0, 137 mM NaCl, 2.7 mM KCl, 1 mM MgCl2, and 0.1 mg/ml BSA, 30 μM Biomol substrate, 200 μM NAD+and 20 ng/μl Sirtuin 1. Incubation condition: 30 min at 30℃.
• Purity: > 80%.
• Stability: >6 months at –80°C.
• Publications:
  Circular and linear: a tale of aptamer selection for the activation of SIRT1 to induce death in cancer cells (2020)

Gene Information

• Gene Name: SIRT1 sirtuin (silent mating type information regulation 2 homolog) 1 (S. cerevisiae) [ Homo sapiens ]
• Synonyms: sirtuin (silent mating type information regulation 2 homolog) 1 (S. cerevisiae); SIR2L1; SIRT1; sirtuin 1; SIR2alpha; sir2-like 1; sirtuin type 1; EC 3.5.1.-; OTTHUMP00000019691; OTTHUMP00000060745; SIR2alpha; hSIR2; hSIRT1; SIR2-like protein 1; sirtuin (silent mating type information regulation 2, S. cerevisiae, homolog)
• Gene ID: 23411
• mRNA Refseq: NM_001142498
• Protein Refseq: NP_001135970
• MIM: 604479
• UniProt ID: Q96EB6
• Chromosome Location: 10q21.3
• Function: NAD binding; NAD+ ADP-ribosyltransferase activity; NAD-dependent histone deacetylase activity; deacetylase activity; histone binding; hydrolase activity, acting on carbon-nitrogen (but not peptide) bonds, in linear amides; identical protein binding; metal ion binding; p53 binding; protein C-terminus binding; protein domain specific binding; transcription corepressor activity; zinc ion binding

Citation

Circular and linear: a tale of aptamer selection for the activation of SIRT1 to induce death in cancer cells

Journal: RSC Advances    PubMed ID: 35516259    Data: 2020/12/21

Authors: Basma Al-Sudani, Abby H. Ragazzon-Smith, Patricia A. Ragazzon

Article Snippet:BEGM BulletKit? was purchased from Lonza.BEGM BulletKit? was purchased from Lonza.. Recombinant human SIRT1-GST-tagged [SIRT1-462H] was purchased from Creative BioMart.. PreScission Protease on-column GSTrap FF and NHS-HP SpinTrap column were purchased from GE HealthCare.PreScission Protease on-column GSTrap FF and NHS-HP SpinTrap column were purchased from GE HealthCare.

Workflow representation of the selection process. In the case of the circular library, the oligonucleotides were phosphorylated and ligated. In the case of the linear library, the oligonucleotides were unmodified. First stage of the cycle starts with the SIRT1 interacting with either library. Unbound sequences were eluted using different NaCl ionic concentration. The bound sequences were then amplified by PCR. In the case of circular library, the sequences were phosphorylated and ligated before entering the second round. The linear library sequences were used from the purification step after the PCR one. Once eight cycles (circular) and twelve (linear) were performed, the sequences were cloned, analysed and studied for biological activity.

Association binding parameters resulting from the analysis of the surface plasmon resonance data for the interaction between the selected aptamers and the SIRT1 enzyme

Fluorescence microscopy analysis of AC3 binding to SIRT1 enzyme and nuclei localisation. AC3-FAM (green-yellow), nuclei (DAPI, blue) and SIRT1 (Texas Red, red). Overlap of images can produced alterations in the colour results such as yellow produced by overlap of red and green, magenta produced by red and blue and cyan produced by green and blue. (A) A549, scale 100 μm. (B) HepG2 (augmentation in Fig. 20, ESI. (C) Caco2, scale 200 μm). (D) U2OS, scale 200 μm. (E) MCF7, scale 20 μm. (F) MCF7 zoomed. (G) MDAMB-468, scale 100 μm. (H) Beas2B, scale 200 μm. Fluorescence intensity was measured by LUMIStar luminometer microplate readers as: green ex. 495 nm, em. 509 nm; blue ex. 359 nm, em. 461 nm and red ex. 596 nm, em. 615 nm. The results represent the mean ± SEM of two different experiments performed twice. Images were taken at a magnification of 100× in a Cytation 3.