14C-labeled acyl-coenzyme A (acetyl-CoA, malonyl-CoA, and succinyl-CoA) was purchased from PerkinElmer (Waltham, MA, USA). Calf thymus histone and unlabeled acyl-coenzyme A were purchased from Sigma-Aldrich (St. Louis, MO, USA). For western blotting, the following antibodies were used: anti-IKKα (Santa Cruz, sc-7218) (CA, USA), anti-HDAC2 (ABR, PA I-861) (Golden, CO, USA), anti-acetyl histone H3 (Millipore, 06-594) (Bedford, MA, USA), anti-E1A-binding protein p300 (p300) (Santa Cruz, sc-585), and anti-CREB-binding protein (CBP) (Abcam, ab3652) (Cambridge, UK).
Acetyl coa
Manufactured by PerkinElmer
Sourced in United States
Acetyl-CoA is a key metabolic intermediate involved in various cellular processes. It serves as a central hub for energy production and biosynthetic pathways. Acetyl-CoA is a fundamental component in the citric acid cycle and plays a critical role in the metabolism of carbohydrates, fats, and proteins.
Automatically generated - may contain errors
Lab products found in correlation
Calf thymus histone, by Merck Group (1 mentions)
Anti acetyl histone h3, by Merck Group (1 mentions)
Malonyl coa, by PerkinElmer (1 mentions)
Anti ikkα, by Santa Cruz Biotechnology (1 mentions)
3h acetyl coa, by PerkinElmer (1 mentions)
Sodium butyrate, by Merck Group (1 mentions)
Anti flag m2 agarose bead, by Merck Group (1 mentions)
Anti ha agarose beads, by Roche (1 mentions)
4 protocols using acetyl coa
1
Histone Acetylation Assay Protocol
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14C-labeled acyl-coenzyme A (acetyl-CoA, malonyl-CoA, and succinyl-CoA) was purchased from PerkinElmer (Waltham, MA, USA). Calf thymus histone and unlabeled acyl-coenzyme A were purchased from Sigma-Aldrich (St. Louis, MO, USA). For western blotting, the following antibodies were used: anti-IKKα (Santa Cruz, sc-7218) (CA, USA), anti-HDAC2 (ABR, PA I-861) (Golden, CO, USA), anti-acetyl histone H3 (Millipore, 06-594) (Bedford, MA, USA), anti-E1A-binding protein p300 (p300) (Santa Cruz, sc-585), and anti-CREB-binding protein (CBP) (Abcam, ab3652) (Cambridge, UK).
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14C-labeled acyl-coenzyme A (acetyl-CoA, malonyl-CoA, and succinyl-CoA) was purchased from PerkinElmer (Waltham, MA, USA). Calf thymus histone and unlabeled acyl-coenzyme A were purchased from Sigma-Aldrich (St. Louis, MO, USA). For western blotting, the following antibodies were used: anti-IKKα (Santa Cruz, sc-7218) (CA, USA), anti-HDAC2 (ABR, PA I-861) (Golden, CO, USA), anti-acetyl histone H3 (Millipore, 06-594) (Bedford, MA, USA), anti-E1A-binding protein p300 (p300) (Santa Cruz, sc-585), and anti-CREB-binding protein (CBP) (Abcam, ab3652) (Cambridge, UK).
2
Histone Acetylation Analysis Protocol
3
Purification and HAT Assay of Protein Complexes
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Purification of protein complexes after co-transfection was performed as previously described (26 (link),27 (link)). Briefly, 293T cells were transfected with each subunit (HA-BRPF1 (wt or mutants), Flag-MOZ, Flag- or HA-ING5 and Flag- or HA-hEaf6) and harvested 48 h later. Flag or HA IP using anti-Flag M2 agarose beads (Sigma) or anti-HA agarose beads (Roche) was performed followed by elution with 3× Flag or HA peptides. HAT assays were performed as described previously (22 (link)). Briefly, native human chromatin and free histones were used to carry out HAT assays in a 15 μl reaction containing 50 mM Tris–HCl pH 8.0, 10% glycerol, 1 mM DTT, 0.1 mM EDTA, 1 mM PMSF, 10 mM sodium butyrate (Sigma) and 1.25 nCi 3H labeled acetyl-CoA (Perkin Elmer Life Sciences). Samples were spotted on P81 membranes (GE Healthcare) for scintillation counting.
4
Radioactive Acetylation Assay for Effectors
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For 14 C-based acetylation assays, 3 µg of the effector (6xHis-HopZ1a, 6xHis-HopZ1a C216A or 6xHis-HopZ1a K289R ) and 5 µg of the substrate (GST-MKK7 or GST-MKK7 K167R ) were incubated in acetylation buffer containing 50 mM HEPES pH 8, 10% Glycerol, 10 mM Sodium butyrate, 1 mM DTT and 1 mM PMSF, with 100 nM inositol hexakisphosphate (IP6 or phytic acid, SIGMA, USA) and 22 nCi Acetyl-CoA (Perkin Elmer, USA). The reaction was incubated for 1 hour at 30ºC, then stopped by adding Laemmli buffer and boiled at 95ºC for 5 minutes. Twenty µl of each sample were separated by SDS-PAGE, and proteins were transferred onto a PVDF membrane.
Acetylation was detected by autoradiography. As a loading control 20 µl of the same samples were separated by SDS-PAGE and stained with Coomassie blue.
Acetylation was detected by autoradiography. As a loading control 20 µl of the same samples were separated by SDS-PAGE and stained with Coomassie blue.
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