fluorogenic acetylated peptide substrate  (BPS Bioscience)

Journal: bioRxiv

Article Title: A small molecule PTER-selective inhibitor reduces food intake and body weight

doi: 10.64898/2026.01.26.701829

Figure Lengend Snippet: (A) Superposition of human HDAC(1-11) structures showing conserved aromatic side chains in active site. Average distance between two side chain is labeled. PDB IDs are 4BKX, 4LXZ, 4A69, 2VQJ, 5EDU, 3C0Y, 1T64 for human HDAC 1-4, 6-8. HDAC5 and 9-11 are AlphaFold-predicted models. The representative SAHA (white) is from an HDAC2 co-crystal structure (PDB ID 4LXZ). (B) Superposition of the top poses of docked PTERi (yellow) in sPTER and docked SAHA in sPTER, and SAHA from an HDAC2 co-crystal structure (PDB ID 4LXZ). Superposition is based on the top-ranked SAHA pose in sPTER and SAHA pose in HDAC2. SAHA poses are not shown for simplicity. Side chains of sPTER are shown in purple, while side chains of HDAC2 are shown in white. (C) Chemical structure of PTERi. (D) Dose-response inhibition of PTER activity by PTERi. (E) Heat map of dose-response inhibition for PTERi against the indicated recombinant enzyme. (F, G) Lineweaver-Burke plot (F) and dose-response inhibition of PTERi (G) in PTER activity assays. For (D-G) , PTER activity (N-acetyltaurine hydrolysis) was measured by quantifying taurine production using 200 ng of purified recombinant mouse PTER (mPTER, panels D-G ) or purified recombinant PTER from the indicated species (G) and 100 µM N-acetyltaurine for 1 h at 37°C. N=3/data point for (D,G) and N=1/data point for (E,F) . (D,G) are shown as mean ± SEM. IC 50 values for were determined from the dose-response curves via nonlinear regression analysis using GraphPad Prism.

Article Snippet: The HDAC assay was performed in 50 μl reaction volume containing the HDAC reaction assay buffer, 600 μg of the liver lysate, 100 μM HDAC substrate peptide Ac-Arg-Gly-Lys-Ac-Glu-AMC (custom synthesized by Elim Biopharm), and 10 μM of indicated inhibitors.

Techniques: Labeling, Inhibition, Activity Assay, Recombinant, Purification

Journal: Current Protocols

Article Title: Epigenetic Histone Deacetylases Activity Assay in the Brain and Peripheral Organ Tissues

doi: 10.1002/cpz1.70143

Figure Lengend Snippet: Biochemical basis of the HDAC activity assay in brain and other tissues. Figure created using BioRender.

Article Snippet: HDAC activity is measured in nuclear lysates using the fluorometric HDAC substrate Boc‐Lys(Ac)‐AMC/Boc‐Lys‐AMC assay (I‐1875 and I‐1880, Bachem), as reported previously (Nian et al., ; Rajendran et al., , ).

Techniques: HDAC Activity Assay

Journal: Current Protocols

Article Title: Epigenetic Histone Deacetylases Activity Assay in the Brain and Peripheral Organ Tissues

doi: 10.1002/cpz1.70143

Figure Lengend Snippet: Illustration of critical steps involved in measuring HDAC activity in the mice tissues. Figure created using BioRender.

Article Snippet: HDAC activity is measured in nuclear lysates using the fluorometric HDAC substrate Boc‐Lys(Ac)‐AMC/Boc‐Lys‐AMC assay (I‐1875 and I‐1880, Bachem), as reported previously (Nian et al., ; Rajendran et al., , ).

Techniques: Activity Assay

Journal: Current Protocols

Article Title: Epigenetic Histone Deacetylases Activity Assay in the Brain and Peripheral Organ Tissues

doi: 10.1002/cpz1.70143

Figure Lengend Snippet: Assay layouts for protein estimation and HDAC activity. ( A ) Representative sample layout of the 96‐well microplate for the BCA assay. ( B ) Representative sample layout of the 96‐well microplate for the HDAC activity assay. ( C ) A typical standard curve generated using the Spectra MAX Gemini XS Plate Reader is shown with standard concentrations of 50, 250, 500, 1000, 1500, and 2000 µg/ml at an excitation wavelength of 360 nm and emission wavelength of 460 nm. Panels A and B were created using BioRender.

Article Snippet: HDAC activity is measured in nuclear lysates using the fluorometric HDAC substrate Boc‐Lys(Ac)‐AMC/Boc‐Lys‐AMC assay (I‐1875 and I‐1880, Bachem), as reported previously (Nian et al., ; Rajendran et al., , ).

Techniques: Activity Assay, BIA-KA, HDAC Activity Assay, Generated

Journal: Current Protocols

Article Title: Epigenetic Histone Deacetylases Activity Assay in the Brain and Peripheral Organ Tissues

doi: 10.1002/cpz1.70143

Figure Lengend Snippet: Quantification of HDAC activity in brain and other tissues. ( A ) Basal HDAC activity in sham brain tissues 7 days after TBI. The spleen is shown to have the highest normalized HDAC activity. ( B ) Percentage HDAC activity normalized to hippocampus levels. ( C and D ) Inhibition of HDAC activity by HDAC inhibitors 24 hr after TBI in the hippocampus of male mice is shown as ( C ) normalized HDAC activity and ( D ) percentage change in HDAC activity. ( E ) Normalized HDAC activity in the cortex and hippocampus of male and female mice 24 hr after TBI. Data were normalized to samples with trichostatin A (TSA) and are presented as normalized HDAC activity. TBI, traumatic brain injury; ROMI, romidepsin; SAHA, vorinostat; SB, sodium butyrate; FEU, fluorescence emission unit. Data represent mean ± S.E.M. ( n = 6 to 8 for each group). In subpanels C and D, * p < .05 versus sham; # p < .05 versus TBI. In subpanel E, * p < .05 versus sham (same sex); # p < .05 versus TBI (across sex); & p < .05 versus sham (across sex). Data were analyzed using one‐way ANOVA followed by Tukey's post hoc test.

Article Snippet: HDAC activity is measured in nuclear lysates using the fluorometric HDAC substrate Boc‐Lys(Ac)‐AMC/Boc‐Lys‐AMC assay (I‐1875 and I‐1880, Bachem), as reported previously (Nian et al., ; Rajendran et al., , ).

Techniques: Activity Assay, Inhibition, Fluorescence