Human HDAC3 Baculovirus-Insect cells Overexpression Lysate

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Human HDAC3 Baculovirus-Insect cells Overexpression Lysate: Product Information

Product Description
This Human HDAC3 overexpression lysate was created in Baculovirus-Insect cells and intented for use as a Western blot (WB) positive control. Purification of HDAC3 protein (Cat: 11511-H20B) from the overexpression lysate was verified.
Expression Host
Baculovirus-Insect cells
Species
Human
Sequence Information
A DNA sequence encoding the full length of human HDAC3 (O15379-1) (Met 1-Ile 428) was fused with the N-terminal polyhistidine-tagged GST tag at the N-terminus.
Molecule Mass
The recombinant human HDAC3/GST chimera consists of 666 amino acids and has a calculated molecular mass of 76.7 kDa. It migrates as an approximately 66 kDa band in SDS-PAGE under reducing conditions.

Human HDAC3 Baculovirus-Insect cells Overexpression Lysate: Usage Guide

Preparation Method
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
Lysis Buffer
Modified RIPA Lysis Buffer: 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1% Sodium deoxycholate, 1mM PMSF.
Recommend Usage
1.  Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube. 2.  Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
Sample Buffer
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Stability & Storage
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
Application
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.

Human HDAC3 Baculovirus-Insect cells Overexpression Lysate: Alternative Names

Human HD3 Overexpression Lysate; Human RPD3 Overexpression Lysate; Human RPD3-2 Overexpression Lysate

HDAC3 Background Information

Histone Deacetylases (HDACs) are a group of enzymes closely related to sirtuins. They catalyze the removal of acetyl groups from lysine residues in histones and non-histone proteins, resulting in transcriptional repression. In general, they do not act autonomously but as components of large multiprotein complexes, such as pRb-E2F and mSin3A, that mediate important transcription regulatory pathways. There are three classes of HDACs; classes 1, 2 and 4, which are closely related Zn2+-dependent enzymes. HDACs are ubiquitously expressed and they can exist in the nucleus or cytosol. Their subcellular localization is effected by protein-protein interactions (for example HDAC-14.3.3 complexes are retained in the cytosol) and by the class to which they belong (class 1 HDACs are predominantly nuclear whilst class 2 HDACs shuttle between the nucleus and cytosol). HDACs have a role in cell growth arrest, differentiation and death and this has led to substantial interest in HDAC inhibitors as possible antineoplastic agents. Histone Deacetylases (HDACs) is Responsible for the deacetylation of lysine residues on the N-terminal part of the core histones (H2A, H2, H3 and H4). Histone deacetylation gives a tag for epigenetic repression and plays an important role in transcriptional regulation, cell cycle progression and developmental events. Histone deacetylases act via the formation of large multiprotein complexes. Probably participates in the regulation of transcription through its binding to the zinc-finger transcription factor YY1; increases YY1 repression activity. Required to repress transcription of the POU1F1 transcription factor. Acts as a molecular chaperone for shuttling phosphorylated NR2C1 to PML bodies for sumoylation.
Full Name
histone deacetylase 3
References
  • Emiliani S, et al. (1998) Characterization of a human RPD3 ortholog, HDAC3. Proc Natl Acad Sci. 95 (6): 2795-800.
  • Dangond F, et al. (1998) Differential display cloning of a novel human histone deacetylase (HDAC3) cDNA from PHA-activated immune cells. Biochem Biophys Res Commun. 242 (3): 648-52.
  • Nicolas E, et al. (2001) The histone deacetylase HDAC3 targets RbAp48 to the retinoblastoma protein. Nucleic Acids Res. 29 (15): 3131-6.
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