Review



native ac‑p53 peptide  (GL Biochem)

 
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
    • 90
      Buy from Supplier

    Structured Review

    GL Biochem native ac‑p53 peptide
    Aspirin acetylates the K408 of SIRT1 and inhibits SIRT1 activity. a) Effects of 2 m m aspirin and 1 m m NAM, 10u m EX527 on SIRT1 activity. The deacetylase activities of recombinant human SIRT1 were measured by monitoring the fluorescence intensity (excitation at 360 nm and emission at 460 nm) using a substrate peptide with one end coupled to a fluorophore and the other end to a quencher. A reaction without NAD + was performed as a negative control. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. b) Representative immunoblot of three independent experiments measuring SIRT1 deacetylase activities. Acetylated <t>p53</t> was incubated with indicated doses of aspirin for 3 h, then p53 acetylation was assessed by immunoblotting with anti‐p53 K382Ac antibody. c) The effects of SIRT1 deacetylation on native acetylated p53 peptide in the assay as indicated were determined by mass spectrometry. Data are representative of three independent experiments. d) D3‐acetylated SIRT1 K408 was identified by a tandem mass spectrometry. HEK293T cells were transfected with FLAG‐SIRT1 plasmids. 24 h after transfection, cells were treated with 2 m m aspirin‐D3. Then the FLAG‐SIRT1 was immunoprecipitated and subject to analysis by a MALDI‐TOF/TOF mass spectrometry. e) WT and mutants of SIRT1 were assayed for SIRT1 steady‐state kinetics. Deacetylase activity of recombinant WT SIRT1, K408R mutant, and K408Q mutant were examined and normalized against protein level. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. f) Deacetylase activity of recombinant WT SIRT1 and site‐specific acetylated SIRT1 was examined and normalized against the protein level; n = 3 biologically independent samples per group, represented as the mean s.e.m.; ** P = 0.0025, two‐tailed Student's t ‐test. g) Representative immunoblot for in vitro aspirin acetylating SIRT1 with or without NAD+. Recombinant human SIRT1 was incubated with indicated doses of aspirin for 3 h with or without the presence of NAD+, then SIRT1 acetylation was assessed by immunoblotting with anti‐Pan‐acetyl‐lysine and anti‐SIRT1‐K408Ac antibody. h) Representative immunoblot of three independent experiments showing acetylated K408‐SIRT1. In vitro deacetylation assay of SIRT1 K408ac by recombinant SIRT1 purified from E. coli were performed and inhibitors(NAM: 500 µ m , Ex527: 10 µ m ) of SIRT1 were added as a control. The purified precipitants were analyzed with anti‐pan‐acetyllysine and anti‐His antibody.
    Native Ac‑P53 Peptide, supplied by GL 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/native+ac-p53+peptide/pmc11109641-254-0-9?v=GL+Biochem
    Average 90 stars, based on 1 article reviews
    native ac‑p53 peptide - by Bioz Stars, 2026-08
    90/100 stars

    Images

    1) Product Images from "Aspirin‐Mediated Acetylation of SIRT1 Maintains Intestinal Immune Homeostasis"

    Article Title: Aspirin‐Mediated Acetylation of SIRT1 Maintains Intestinal Immune Homeostasis

    Journal: Advanced Science

    doi: 10.1002/advs.202306378

    Aspirin acetylates the K408 of SIRT1 and inhibits SIRT1 activity. a) Effects of 2 m m aspirin and 1 m m NAM, 10u m EX527 on SIRT1 activity. The deacetylase activities of recombinant human SIRT1 were measured by monitoring the fluorescence intensity (excitation at 360 nm and emission at 460 nm) using a substrate peptide with one end coupled to a fluorophore and the other end to a quencher. A reaction without NAD + was performed as a negative control. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. b) Representative immunoblot of three independent experiments measuring SIRT1 deacetylase activities. Acetylated p53 was incubated with indicated doses of aspirin for 3 h, then p53 acetylation was assessed by immunoblotting with anti‐p53 K382Ac antibody. c) The effects of SIRT1 deacetylation on native acetylated p53 peptide in the assay as indicated were determined by mass spectrometry. Data are representative of three independent experiments. d) D3‐acetylated SIRT1 K408 was identified by a tandem mass spectrometry. HEK293T cells were transfected with FLAG‐SIRT1 plasmids. 24 h after transfection, cells were treated with 2 m m aspirin‐D3. Then the FLAG‐SIRT1 was immunoprecipitated and subject to analysis by a MALDI‐TOF/TOF mass spectrometry. e) WT and mutants of SIRT1 were assayed for SIRT1 steady‐state kinetics. Deacetylase activity of recombinant WT SIRT1, K408R mutant, and K408Q mutant were examined and normalized against protein level. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. f) Deacetylase activity of recombinant WT SIRT1 and site‐specific acetylated SIRT1 was examined and normalized against the protein level; n = 3 biologically independent samples per group, represented as the mean s.e.m.; ** P = 0.0025, two‐tailed Student's t ‐test. g) Representative immunoblot for in vitro aspirin acetylating SIRT1 with or without NAD+. Recombinant human SIRT1 was incubated with indicated doses of aspirin for 3 h with or without the presence of NAD+, then SIRT1 acetylation was assessed by immunoblotting with anti‐Pan‐acetyl‐lysine and anti‐SIRT1‐K408Ac antibody. h) Representative immunoblot of three independent experiments showing acetylated K408‐SIRT1. In vitro deacetylation assay of SIRT1 K408ac by recombinant SIRT1 purified from E. coli were performed and inhibitors(NAM: 500 µ m , Ex527: 10 µ m ) of SIRT1 were added as a control. The purified precipitants were analyzed with anti‐pan‐acetyllysine and anti‐His antibody.
    Figure Legend Snippet: Aspirin acetylates the K408 of SIRT1 and inhibits SIRT1 activity. a) Effects of 2 m m aspirin and 1 m m NAM, 10u m EX527 on SIRT1 activity. The deacetylase activities of recombinant human SIRT1 were measured by monitoring the fluorescence intensity (excitation at 360 nm and emission at 460 nm) using a substrate peptide with one end coupled to a fluorophore and the other end to a quencher. A reaction without NAD + was performed as a negative control. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. b) Representative immunoblot of three independent experiments measuring SIRT1 deacetylase activities. Acetylated p53 was incubated with indicated doses of aspirin for 3 h, then p53 acetylation was assessed by immunoblotting with anti‐p53 K382Ac antibody. c) The effects of SIRT1 deacetylation on native acetylated p53 peptide in the assay as indicated were determined by mass spectrometry. Data are representative of three independent experiments. d) D3‐acetylated SIRT1 K408 was identified by a tandem mass spectrometry. HEK293T cells were transfected with FLAG‐SIRT1 plasmids. 24 h after transfection, cells were treated with 2 m m aspirin‐D3. Then the FLAG‐SIRT1 was immunoprecipitated and subject to analysis by a MALDI‐TOF/TOF mass spectrometry. e) WT and mutants of SIRT1 were assayed for SIRT1 steady‐state kinetics. Deacetylase activity of recombinant WT SIRT1, K408R mutant, and K408Q mutant were examined and normalized against protein level. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. f) Deacetylase activity of recombinant WT SIRT1 and site‐specific acetylated SIRT1 was examined and normalized against the protein level; n = 3 biologically independent samples per group, represented as the mean s.e.m.; ** P = 0.0025, two‐tailed Student's t ‐test. g) Representative immunoblot for in vitro aspirin acetylating SIRT1 with or without NAD+. Recombinant human SIRT1 was incubated with indicated doses of aspirin for 3 h with or without the presence of NAD+, then SIRT1 acetylation was assessed by immunoblotting with anti‐Pan‐acetyl‐lysine and anti‐SIRT1‐K408Ac antibody. h) Representative immunoblot of three independent experiments showing acetylated K408‐SIRT1. In vitro deacetylation assay of SIRT1 K408ac by recombinant SIRT1 purified from E. coli were performed and inhibitors(NAM: 500 µ m , Ex527: 10 µ m ) of SIRT1 were added as a control. The purified precipitants were analyzed with anti‐pan‐acetyllysine and anti‐His antibody.

    Techniques Used: Activity Assay, Histone Deacetylase Assay, Recombinant, Fluorescence, Negative Control, Western Blot, Incubation, Mass Spectrometry, Transfection, Immunoprecipitation, Mutagenesis, Two Tailed Test, In Vitro, Purification, Control

    Inactivation of acetylated K408 of SIRT1 due to impaired substrates binding affinity. a) ITC measurements of binding affinities of NAD+ to the recombinant WT SIRT1 and K408Q mutant. b) Titration curves for determination of Km values of recombinant WT SIRT1 and K408Q mutant toward the indicated fluorogenic substrates and NAD+. A summary of measured Michaelis–Menten kinetic parameters for the indicated fluorogenic substrates and NAD+. c,d) Overview of the SIRT1 structure highlighting the position of K408 (in cyan‐colored spheres). The model is adapted from crystal structures of SIRT1 bound with either ADPR (PDB: 4KXQ) or a peptide substrate p53‐AMC (PDB: 5BTR). The structured loop connecting the Zn 2+ ‐binding domain and the NAD‐binding domain is shown in purple. e) A close‐up view of the interactions between K408 and the connecting loop.
    Figure Legend Snippet: Inactivation of acetylated K408 of SIRT1 due to impaired substrates binding affinity. a) ITC measurements of binding affinities of NAD+ to the recombinant WT SIRT1 and K408Q mutant. b) Titration curves for determination of Km values of recombinant WT SIRT1 and K408Q mutant toward the indicated fluorogenic substrates and NAD+. A summary of measured Michaelis–Menten kinetic parameters for the indicated fluorogenic substrates and NAD+. c,d) Overview of the SIRT1 structure highlighting the position of K408 (in cyan‐colored spheres). The model is adapted from crystal structures of SIRT1 bound with either ADPR (PDB: 4KXQ) or a peptide substrate p53‐AMC (PDB: 5BTR). The structured loop connecting the Zn 2+ ‐binding domain and the NAD‐binding domain is shown in purple. e) A close‐up view of the interactions between K408 and the connecting loop.

    Techniques Used: Binding Assay, Recombinant, Mutagenesis, Titration

    SIRT1 is required for Aspirin to induce apoptosis. a) Representative immunoblot of three independent experiments showing the analysis of SIRT1 and its substrates p53ac in HCT116 cells treated with different doses of Aspirin for 24 h. p53 acetylation was assessed after p53 immunoprecipitation. b) Representative immunoblot of three independent experiments showing the acetylated levels of downstream targets of p53. HCT116 cells were transfected with control and SIRT1 siRNAs. Cells were treated as described before and western blot analysis of SIRT1, BAX, PUMA in cell extracts, and p53 acetylation after p53 immunoprecipitation. c) HCT116(WT) and HCT116(p53‐/‐) cells were treated with DMSO or 2 m m aspirin, and apoptosis was measured by FACS. Overexpression of SIRT1 suppresses apoptosis but only in the presence of p53. Error bars represent s.d., n = 3. d) HCT116(WT) and HCT116(SIRT1 KO) cells were treated with DMSO or 2 m m aspirin, and apoptosis was measured by FACS. Error bars represent s.d., n = 3. e) Cell growth of HCT116(WT) and HCT116(SIRT1 KO) cells incubated with DMSO or 2 m m aspirin. The cell numbers were detected by the CCK8 kit. f) HCT116(WT) and HCT116(SIRT1‐KO) cells were treated with DMSO or 2 m m aspirin to perform a xenograft assay in nude mice. Tumors were photographed and weighed. Error bars represent s.d., n = 6.
    Figure Legend Snippet: SIRT1 is required for Aspirin to induce apoptosis. a) Representative immunoblot of three independent experiments showing the analysis of SIRT1 and its substrates p53ac in HCT116 cells treated with different doses of Aspirin for 24 h. p53 acetylation was assessed after p53 immunoprecipitation. b) Representative immunoblot of three independent experiments showing the acetylated levels of downstream targets of p53. HCT116 cells were transfected with control and SIRT1 siRNAs. Cells were treated as described before and western blot analysis of SIRT1, BAX, PUMA in cell extracts, and p53 acetylation after p53 immunoprecipitation. c) HCT116(WT) and HCT116(p53‐/‐) cells were treated with DMSO or 2 m m aspirin, and apoptosis was measured by FACS. Overexpression of SIRT1 suppresses apoptosis but only in the presence of p53. Error bars represent s.d., n = 3. d) HCT116(WT) and HCT116(SIRT1 KO) cells were treated with DMSO or 2 m m aspirin, and apoptosis was measured by FACS. Error bars represent s.d., n = 3. e) Cell growth of HCT116(WT) and HCT116(SIRT1 KO) cells incubated with DMSO or 2 m m aspirin. The cell numbers were detected by the CCK8 kit. f) HCT116(WT) and HCT116(SIRT1‐KO) cells were treated with DMSO or 2 m m aspirin to perform a xenograft assay in nude mice. Tumors were photographed and weighed. Error bars represent s.d., n = 6.

    Techniques Used: Western Blot, Immunoprecipitation, Transfection, Control, Over Expression, Incubation, Xenograft Assay



    Similar Products

    native ac‑p53 peptide  (GL Biochem)
    90
    GL Biochem native ac‑p53 peptide
    Aspirin acetylates the K408 of SIRT1 and inhibits SIRT1 activity. a) Effects of 2 m m aspirin and 1 m m NAM, 10u m EX527 on SIRT1 activity. The deacetylase activities of recombinant human SIRT1 were measured by monitoring the fluorescence intensity (excitation at 360 nm and emission at 460 nm) using a substrate peptide with one end coupled to a fluorophore and the other end to a quencher. A reaction without NAD + was performed as a negative control. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. b) Representative immunoblot of three independent experiments measuring SIRT1 deacetylase activities. Acetylated <t>p53</t> was incubated with indicated doses of aspirin for 3 h, then p53 acetylation was assessed by immunoblotting with anti‐p53 K382Ac antibody. c) The effects of SIRT1 deacetylation on native acetylated p53 peptide in the assay as indicated were determined by mass spectrometry. Data are representative of three independent experiments. d) D3‐acetylated SIRT1 K408 was identified by a tandem mass spectrometry. HEK293T cells were transfected with FLAG‐SIRT1 plasmids. 24 h after transfection, cells were treated with 2 m m aspirin‐D3. Then the FLAG‐SIRT1 was immunoprecipitated and subject to analysis by a MALDI‐TOF/TOF mass spectrometry. e) WT and mutants of SIRT1 were assayed for SIRT1 steady‐state kinetics. Deacetylase activity of recombinant WT SIRT1, K408R mutant, and K408Q mutant were examined and normalized against protein level. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. f) Deacetylase activity of recombinant WT SIRT1 and site‐specific acetylated SIRT1 was examined and normalized against the protein level; n = 3 biologically independent samples per group, represented as the mean s.e.m.; ** P = 0.0025, two‐tailed Student's t ‐test. g) Representative immunoblot for in vitro aspirin acetylating SIRT1 with or without NAD+. Recombinant human SIRT1 was incubated with indicated doses of aspirin for 3 h with or without the presence of NAD+, then SIRT1 acetylation was assessed by immunoblotting with anti‐Pan‐acetyl‐lysine and anti‐SIRT1‐K408Ac antibody. h) Representative immunoblot of three independent experiments showing acetylated K408‐SIRT1. In vitro deacetylation assay of SIRT1 K408ac by recombinant SIRT1 purified from E. coli were performed and inhibitors(NAM: 500 µ m , Ex527: 10 µ m ) of SIRT1 were added as a control. The purified precipitants were analyzed with anti‐pan‐acetyllysine and anti‐His antibody.
    Native Ac‑P53 Peptide, supplied by GL 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/native+ac-p53+peptide/pmc11109641-254-0-9?v=GL+Biochem
    Average 90 stars, based on 1 article reviews
    native ac‑p53 peptide - by Bioz Stars, 2026-08
    90/100 stars
    		  Buy from Supplier
    native ac-p53 peptide  (GL Biochem)
    90
    GL Biochem native ac-p53 peptide
    Aspirin acetylates the K408 of SIRT1 and inhibits SIRT1 activity. a) Effects of 2 m m aspirin and 1 m m NAM, 10u m EX527 on SIRT1 activity. The deacetylase activities of recombinant human SIRT1 were measured by monitoring the fluorescence intensity (excitation at 360 nm and emission at 460 nm) using a substrate peptide with one end coupled to a fluorophore and the other end to a quencher. A reaction without NAD + was performed as a negative control. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. b) Representative immunoblot of three independent experiments measuring SIRT1 deacetylase activities. Acetylated <t>p53</t> was incubated with indicated doses of aspirin for 3 h, then p53 acetylation was assessed by immunoblotting with anti‐p53 K382Ac antibody. c) The effects of SIRT1 deacetylation on native acetylated p53 peptide in the assay as indicated were determined by mass spectrometry. Data are representative of three independent experiments. d) D3‐acetylated SIRT1 K408 was identified by a tandem mass spectrometry. HEK293T cells were transfected with FLAG‐SIRT1 plasmids. 24 h after transfection, cells were treated with 2 m m aspirin‐D3. Then the FLAG‐SIRT1 was immunoprecipitated and subject to analysis by a MALDI‐TOF/TOF mass spectrometry. e) WT and mutants of SIRT1 were assayed for SIRT1 steady‐state kinetics. Deacetylase activity of recombinant WT SIRT1, K408R mutant, and K408Q mutant were examined and normalized against protein level. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. f) Deacetylase activity of recombinant WT SIRT1 and site‐specific acetylated SIRT1 was examined and normalized against the protein level; n = 3 biologically independent samples per group, represented as the mean s.e.m.; ** P = 0.0025, two‐tailed Student's t ‐test. g) Representative immunoblot for in vitro aspirin acetylating SIRT1 with or without NAD+. Recombinant human SIRT1 was incubated with indicated doses of aspirin for 3 h with or without the presence of NAD+, then SIRT1 acetylation was assessed by immunoblotting with anti‐Pan‐acetyl‐lysine and anti‐SIRT1‐K408Ac antibody. h) Representative immunoblot of three independent experiments showing acetylated K408‐SIRT1. In vitro deacetylation assay of SIRT1 K408ac by recombinant SIRT1 purified from E. coli were performed and inhibitors(NAM: 500 µ m , Ex527: 10 µ m ) of SIRT1 were added as a control. The purified precipitants were analyzed with anti‐pan‐acetyllysine and anti‐His antibody.
    Native Ac P53 Peptide, supplied by GL 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/native+ac-p53+peptide/pm38482749-362-11-21?v=GL+Biochem
    Average 90 stars, based on 1 article reviews
    native ac-p53 peptide - by Bioz Stars, 2026-08
    90/100 stars
    		  Buy from Supplier

    Image Search Results


    Aspirin acetylates the K408 of SIRT1 and inhibits SIRT1 activity. a) Effects of 2 m m aspirin and 1 m m NAM, 10u m EX527 on SIRT1 activity. The deacetylase activities of recombinant human SIRT1 were measured by monitoring the fluorescence intensity (excitation at 360 nm and emission at 460 nm) using a substrate peptide with one end coupled to a fluorophore and the other end to a quencher. A reaction without NAD + was performed as a negative control. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. b) Representative immunoblot of three independent experiments measuring SIRT1 deacetylase activities. Acetylated p53 was incubated with indicated doses of aspirin for 3 h, then p53 acetylation was assessed by immunoblotting with anti‐p53 K382Ac antibody. c) The effects of SIRT1 deacetylation on native acetylated p53 peptide in the assay as indicated were determined by mass spectrometry. Data are representative of three independent experiments. d) D3‐acetylated SIRT1 K408 was identified by a tandem mass spectrometry. HEK293T cells were transfected with FLAG‐SIRT1 plasmids. 24 h after transfection, cells were treated with 2 m m aspirin‐D3. Then the FLAG‐SIRT1 was immunoprecipitated and subject to analysis by a MALDI‐TOF/TOF mass spectrometry. e) WT and mutants of SIRT1 were assayed for SIRT1 steady‐state kinetics. Deacetylase activity of recombinant WT SIRT1, K408R mutant, and K408Q mutant were examined and normalized against protein level. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. f) Deacetylase activity of recombinant WT SIRT1 and site‐specific acetylated SIRT1 was examined and normalized against the protein level; n = 3 biologically independent samples per group, represented as the mean s.e.m.; ** P = 0.0025, two‐tailed Student's t ‐test. g) Representative immunoblot for in vitro aspirin acetylating SIRT1 with or without NAD+. Recombinant human SIRT1 was incubated with indicated doses of aspirin for 3 h with or without the presence of NAD+, then SIRT1 acetylation was assessed by immunoblotting with anti‐Pan‐acetyl‐lysine and anti‐SIRT1‐K408Ac antibody. h) Representative immunoblot of three independent experiments showing acetylated K408‐SIRT1. In vitro deacetylation assay of SIRT1 K408ac by recombinant SIRT1 purified from E. coli were performed and inhibitors(NAM: 500 µ m , Ex527: 10 µ m ) of SIRT1 were added as a control. The purified precipitants were analyzed with anti‐pan‐acetyllysine and anti‐His antibody.

    Journal: Advanced Science

    Article Title: Aspirin‐Mediated Acetylation of SIRT1 Maintains Intestinal Immune Homeostasis

    doi: 10.1002/advs.202306378

    Figure Lengend Snippet: Aspirin acetylates the K408 of SIRT1 and inhibits SIRT1 activity. a) Effects of 2 m m aspirin and 1 m m NAM, 10u m EX527 on SIRT1 activity. The deacetylase activities of recombinant human SIRT1 were measured by monitoring the fluorescence intensity (excitation at 360 nm and emission at 460 nm) using a substrate peptide with one end coupled to a fluorophore and the other end to a quencher. A reaction without NAD + was performed as a negative control. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. b) Representative immunoblot of three independent experiments measuring SIRT1 deacetylase activities. Acetylated p53 was incubated with indicated doses of aspirin for 3 h, then p53 acetylation was assessed by immunoblotting with anti‐p53 K382Ac antibody. c) The effects of SIRT1 deacetylation on native acetylated p53 peptide in the assay as indicated were determined by mass spectrometry. Data are representative of three independent experiments. d) D3‐acetylated SIRT1 K408 was identified by a tandem mass spectrometry. HEK293T cells were transfected with FLAG‐SIRT1 plasmids. 24 h after transfection, cells were treated with 2 m m aspirin‐D3. Then the FLAG‐SIRT1 was immunoprecipitated and subject to analysis by a MALDI‐TOF/TOF mass spectrometry. e) WT and mutants of SIRT1 were assayed for SIRT1 steady‐state kinetics. Deacetylase activity of recombinant WT SIRT1, K408R mutant, and K408Q mutant were examined and normalized against protein level. Data are presented as mean ± s.d., n = 3 wells, from three independent experiments. f) Deacetylase activity of recombinant WT SIRT1 and site‐specific acetylated SIRT1 was examined and normalized against the protein level; n = 3 biologically independent samples per group, represented as the mean s.e.m.; ** P = 0.0025, two‐tailed Student's t ‐test. g) Representative immunoblot for in vitro aspirin acetylating SIRT1 with or without NAD+. Recombinant human SIRT1 was incubated with indicated doses of aspirin for 3 h with or without the presence of NAD+, then SIRT1 acetylation was assessed by immunoblotting with anti‐Pan‐acetyl‐lysine and anti‐SIRT1‐K408Ac antibody. h) Representative immunoblot of three independent experiments showing acetylated K408‐SIRT1. In vitro deacetylation assay of SIRT1 K408ac by recombinant SIRT1 purified from E. coli were performed and inhibitors(NAM: 500 µ m , Ex527: 10 µ m ) of SIRT1 were added as a control. The purified precipitants were analyzed with anti‐pan‐acetyllysine and anti‐His antibody.

    Article Snippet: Native Ac‐p53 peptide (Ac‐Lys‐Lys‐Gly‐Gln‐Ser‐Thr‐Ser‐Arg‐His‐Lys‐(Ac‐Lys)‐Leu‐Met‐Phe‐Lys‐Thr‐Glu‐Gly‐NH 2 ) was synthesized from GL Biochem.

    Techniques: Activity Assay, Histone Deacetylase Assay, Recombinant, Fluorescence, Negative Control, Western Blot, Incubation, Mass Spectrometry, Transfection, Immunoprecipitation, Mutagenesis, Two Tailed Test, In Vitro, Purification, Control

    Inactivation of acetylated K408 of SIRT1 due to impaired substrates binding affinity. a) ITC measurements of binding affinities of NAD+ to the recombinant WT SIRT1 and K408Q mutant. b) Titration curves for determination of Km values of recombinant WT SIRT1 and K408Q mutant toward the indicated fluorogenic substrates and NAD+. A summary of measured Michaelis–Menten kinetic parameters for the indicated fluorogenic substrates and NAD+. c,d) Overview of the SIRT1 structure highlighting the position of K408 (in cyan‐colored spheres). The model is adapted from crystal structures of SIRT1 bound with either ADPR (PDB: 4KXQ) or a peptide substrate p53‐AMC (PDB: 5BTR). The structured loop connecting the Zn 2+ ‐binding domain and the NAD‐binding domain is shown in purple. e) A close‐up view of the interactions between K408 and the connecting loop.

    Journal: Advanced Science

    Article Title: Aspirin‐Mediated Acetylation of SIRT1 Maintains Intestinal Immune Homeostasis

    doi: 10.1002/advs.202306378

    Figure Lengend Snippet: Inactivation of acetylated K408 of SIRT1 due to impaired substrates binding affinity. a) ITC measurements of binding affinities of NAD+ to the recombinant WT SIRT1 and K408Q mutant. b) Titration curves for determination of Km values of recombinant WT SIRT1 and K408Q mutant toward the indicated fluorogenic substrates and NAD+. A summary of measured Michaelis–Menten kinetic parameters for the indicated fluorogenic substrates and NAD+. c,d) Overview of the SIRT1 structure highlighting the position of K408 (in cyan‐colored spheres). The model is adapted from crystal structures of SIRT1 bound with either ADPR (PDB: 4KXQ) or a peptide substrate p53‐AMC (PDB: 5BTR). The structured loop connecting the Zn 2+ ‐binding domain and the NAD‐binding domain is shown in purple. e) A close‐up view of the interactions between K408 and the connecting loop.

    Article Snippet: Native Ac‐p53 peptide (Ac‐Lys‐Lys‐Gly‐Gln‐Ser‐Thr‐Ser‐Arg‐His‐Lys‐(Ac‐Lys)‐Leu‐Met‐Phe‐Lys‐Thr‐Glu‐Gly‐NH 2 ) was synthesized from GL Biochem.

    Techniques: Binding Assay, Recombinant, Mutagenesis, Titration

    SIRT1 is required for Aspirin to induce apoptosis. a) Representative immunoblot of three independent experiments showing the analysis of SIRT1 and its substrates p53ac in HCT116 cells treated with different doses of Aspirin for 24 h. p53 acetylation was assessed after p53 immunoprecipitation. b) Representative immunoblot of three independent experiments showing the acetylated levels of downstream targets of p53. HCT116 cells were transfected with control and SIRT1 siRNAs. Cells were treated as described before and western blot analysis of SIRT1, BAX, PUMA in cell extracts, and p53 acetylation after p53 immunoprecipitation. c) HCT116(WT) and HCT116(p53‐/‐) cells were treated with DMSO or 2 m m aspirin, and apoptosis was measured by FACS. Overexpression of SIRT1 suppresses apoptosis but only in the presence of p53. Error bars represent s.d., n = 3. d) HCT116(WT) and HCT116(SIRT1 KO) cells were treated with DMSO or 2 m m aspirin, and apoptosis was measured by FACS. Error bars represent s.d., n = 3. e) Cell growth of HCT116(WT) and HCT116(SIRT1 KO) cells incubated with DMSO or 2 m m aspirin. The cell numbers were detected by the CCK8 kit. f) HCT116(WT) and HCT116(SIRT1‐KO) cells were treated with DMSO or 2 m m aspirin to perform a xenograft assay in nude mice. Tumors were photographed and weighed. Error bars represent s.d., n = 6.

    Journal: Advanced Science

    Article Title: Aspirin‐Mediated Acetylation of SIRT1 Maintains Intestinal Immune Homeostasis

    doi: 10.1002/advs.202306378

    Figure Lengend Snippet: SIRT1 is required for Aspirin to induce apoptosis. a) Representative immunoblot of three independent experiments showing the analysis of SIRT1 and its substrates p53ac in HCT116 cells treated with different doses of Aspirin for 24 h. p53 acetylation was assessed after p53 immunoprecipitation. b) Representative immunoblot of three independent experiments showing the acetylated levels of downstream targets of p53. HCT116 cells were transfected with control and SIRT1 siRNAs. Cells were treated as described before and western blot analysis of SIRT1, BAX, PUMA in cell extracts, and p53 acetylation after p53 immunoprecipitation. c) HCT116(WT) and HCT116(p53‐/‐) cells were treated with DMSO or 2 m m aspirin, and apoptosis was measured by FACS. Overexpression of SIRT1 suppresses apoptosis but only in the presence of p53. Error bars represent s.d., n = 3. d) HCT116(WT) and HCT116(SIRT1 KO) cells were treated with DMSO or 2 m m aspirin, and apoptosis was measured by FACS. Error bars represent s.d., n = 3. e) Cell growth of HCT116(WT) and HCT116(SIRT1 KO) cells incubated with DMSO or 2 m m aspirin. The cell numbers were detected by the CCK8 kit. f) HCT116(WT) and HCT116(SIRT1‐KO) cells were treated with DMSO or 2 m m aspirin to perform a xenograft assay in nude mice. Tumors were photographed and weighed. Error bars represent s.d., n = 6.

    Article Snippet: Native Ac‐p53 peptide (Ac‐Lys‐Lys‐Gly‐Gln‐Ser‐Thr‐Ser‐Arg‐His‐Lys‐(Ac‐Lys)‐Leu‐Met‐Phe‐Lys‐Thr‐Glu‐Gly‐NH 2 ) was synthesized from GL Biochem.

    Techniques: Western Blot, Immunoprecipitation, Transfection, Control, Over Expression, Incubation, Xenograft Assay