Enzymatic deacetylation of histone (original) (raw)
Related papers
Methods, 2003
Dynamic histone acetylation has a role in chromatin remodeling and in the regulation of transcription. Histone deacetylases (HDACs) and histone acetyltransferases (HATs) catalyze reversible histone acetylation. HATs and HDACs exist as multiprotein complexes that have coactivator and corepressor activities, respectively. The steady-state level of acetylation at a chromatin site is determined by the local net activities of these enzymes. Here we describe methods to isolate different subcellular fractions (cytosol, nuclei, tightly bound nuclear, loosely bound nuclear, immunoprecipitated multiprotein complexes, and nuclear matrix) to determine the subcellular distribution of HAT and HDAC activities. Procedures to assay the activities of these enzymes and to measure the kinetics of histone acetylation and deacetylation are presented. Ó 2003 Published by Elsevier Science (USA).
Properties of chicken erythrocyte histone deacetylase associated with the nuclear matrix
The Biochemical journal, 1996
Histone H2B is deacetylated more rapidly than H3 and H4 in chicken immature erythrocytes. Histone deacetylase from chicken immature erythrocytes was partially purified, and the histone specificities of the multiple histone deacetylase forms were determined. Ion-exchange (Q-Sepharose) and gel-exclusion (Superdex 200) chromatography of extracts from erythrocyte nuclei showed two forms (HD1 and HD2) of histone deacetylase. HD1, with a molecular mass of about 55 kDa, preferred free H3-H4 relative to H2A-H2B, while HD2, with a molecular mass of approx. 220 kDa, had a slight preference for H3-H4. HD1 and HD2 differed in pH- and ion-strength-dependence. HD2 dissociated into HD1 when treated with 1.6 M NaCl or when applied to a Q-Sepharose column. The enzymic properties of nuclear-matrix-bound histone deacetylase showed a striking difference from that of HD1 and HD2, particularly in its strong preference for H2A-H2B. Treatment of the nuclear matrix with 1.6 M NaCl and 1% 2-mercaptoethanol s...
Histone Deacetylases: Purification of the Enzymes, Substrates, and Assay Conditions
Methods in Enzymology, 2003
Histone deacetylase (HDAC) enzymes catalyze the removal of acetyl groups from the epsilon amino group of conserved lysine residues in the amino terminal tail domains of histones. This modification strengthens histone-DNA interactions and potentially generates specific docking surfaces for proteins that regulate chromatin folding and/or transcription. In addition to their crucial roles in chromatin regulation, HDACs are involved in many different cellular processes such as cell cycle control and differentiation. Although the discovery of HDACs dates back to 1969, 1 research in this area of histone modification was stagnant for a long time because attempts 1 A. Inoue and D. Fujimoto, Biochem. Biophys. Res. Commun. 36, 146 (1969).
Comparative studies on highly metabolically active histone acetylation
Biochimica Et Biophysica Acta (bba) - Nucleic Acids and Protein Synthesis, 1979
Histone acetate is hydrolyzed rapidly in logarithmically dividing hepatoma tissue culture cells (Jackson, V., Shires, A., Chalkley, R. and Granner, D.K. (1975) J. Biol. Chem. 250, 4856--4863). The phenomenon has been analyzed further in hepatoma tissue culture cells at various stages of the cell cycle, in stationary phase, and in the presence of actinomycin D. We also investigated the phenomenon in Tetrahymena pyriformis macronuclei, bovine thymocytes, and human foreskin fibroblasts. The data suggest that this highly metabolically active histone acetylation while altered in mitotic cells, is independent of the overall rate of cell division, and is only slightly sensitive to actinomycin D. Finally, we conclude that the same general phenomenon is found in both cancerous and normal cells and is apparently common to cells from various stages of the evolutionary scale.
Biochimica et biophysica acta, 1979
Nuclei from hepatoma tissue culture (HTC) cells were isolated by standard methods and incubated in media commonly used for nuclease digestions (DNAase I and micrococcal nuclease) and for in vitro RNA synthesis. During the incubation, histones can be deacetylated from both control cells and cells treated with 6 mM sodium butyrate to enhance the levels of histone acetylation. Deacetylation of histone is much more apparent in nuclei isolated from sodium butyrate-treated cells. Inclusion of 6 mM sodium butyrate in the incubation medium effectively inhibits the endogenous deacetylase activity acting on histones H3 and H4, whereas sodium acetate at the same concentration has very little inhibitory effect.
Two species of histone acetyltransferase in rat liver nuclei
Journal of Biological Chemistry
Subcellular localization of histone acetyltransferase was studied in rat liver cells. Two histone acetyltransferases, designated NI and NII, were identified in the nuclear fraction, and an additional two acetyltransferases, termed CI and CII, were separated from the cytoplasmic fraction. These acetyltransferases exhibited different substrate specificities toward free and nucleosomal histones. The enzymes NI and NII represented major histone acetyltransferase activities in rat liver nuclei, and they were further differentiated by DNA-binding properties, subnuclear localization, and reaction kinetics. While the NI enzyme exhibited an intersecting initial velocity kinetic, the NII enzyme followed a ping-pong initial velocity pattern. These results show the multiple occurrence of histone acetyltransferases in nuclear and cytoplasmic fractions, events which may reflect the complexities of histone acetylation.
Modification of histones immediately following synthesis
Archives of Biochemistry and Biophysics, 1979
The acetylation of newly synthesized histones has been studied by exploiting the ability of sodium butyrate to inhibit deacetylation. Upon arrival in the nucleus a significant fraction of histones H2a, H2b, and H3 is in the parental unmodified form. In contrast histone H4 first appears primarily in the diacetylated form. During an ensuing 135 min the diacetylated H4 is slowly further modified.
A comparative study of histone deacetylases of plant, fungal and vertebrate cells
Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology, 1996
The enzymatic equilibrium of reversible core histone acetylation is maintained by two enzyme activities, histone acetyltransferase and histone deacetylase (HD). These enzyme activities exist as multiple enzyme forms. The present report describes methods to extract different HD-forms from three., organisms, germinating maize embryos, the myxomycete Physarum polycephalum, and chicken red blood cells; it provides data on the chromatographic separation and partial purification of HD-forms. In germinating maize embryos three HDs (HD1-A, HD1-B, HD2) can be discriminated; HD1-A, HDI-B, and HD2 were characterized in terms of their dependence on pH, temperature and various ions. as well as kinetic parameters (k M for core histones) and inhibition by various compounds. The same parameters were investigated for the corresponding enzymes of Physarum polycephalum, and mature and immature chicken erythrocytes.
Histone deacetylase is a component of the internal nuclear matrix
Journal of Biological Chemistry
In chicken immature erythrocytes, approximately 4% of the modifiable histone lysine sites participate in active acetylation. There are two categories of actively acetylated histone H4. Although both are acetylated at the same rate (t1/2 = 12 min), one is acetylated to the tetraacetylated form and is rapidly deacetylated (class 1), and the other is acetylated to mono- and diacetylated forms and is slowly deacetylated (class 2). We show that the chromatin distribution of the class 1 labeled tetraacetylated H4 species paralleled that of the transcriptionally active DNA sequences. For example, the chromatin fragments of the insoluble nuclear material contained 76% of the active DNA and 74% of the labeled tetraacetylated H4. Class 2 labeled acetylated H4 species were found in repressed chromatin and were enriched in active/competent gene-enriched chromatin fragments. The majority of the histone deacetylase activity (75-80%) was located with the insoluble residual nuclear material. Furthe...