An activity gel assay detects a single, catalytically active histone acetyltransferase subunit in Tetrahymena macronuclei

Proc Natl Acad Sci U S A. 1995 Jul 3;92(14):6364-8. doi: 10.1073/pnas.92.14.6364.

Abstract

Macronuclei of the ciliated protozoan Tetrahymena thermophila possess a histone acetyltransferase activity closely associated with transcription-related histone acetylation. Nothing definitive is known concerning the polypeptide composition of this activity in Tetrahymena or any comparable activity from any cellular source. An acetyltransferase activity gel assay was developed which identifies a catalytically active subunit of this enzyme in Tetrahymena. This activity gel assay detects a single polypeptide of 55 kDa (p55) in crude macronuclear extracts, as well as in column-purified fractions, which incorporates [3H]acetate from [3H]acetyl-CoA into core histone substrates polymerized directly into SDS polyacrylamide gels. p55 copurifies precisely with acetyltransferase activity through all chromatographic steps examined, including reverse-phase HPLC. Gel-filtration chromatography of this activity indicates a molecular mass of 220 kDa, suggesting that the native enzyme may consist of four identical subunits of 55 kDa. Furthermore, p55 is tightly associated with di- and greater polynucleosomes and therefore may be defined as a component of histone acetyltransferase type A--i.e., chromatin associated.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Acetyltransferases / analysis
  • Acetyltransferases / isolation & purification
  • Acetyltransferases / metabolism*
  • Animals
  • Cell Fractionation
  • Cell Nucleus / enzymology*
  • Chromatin / enzymology*
  • Chromatography
  • Chromatography, Gel
  • Chromatography, High Pressure Liquid
  • Chromatography, Ion Exchange
  • Durapatite
  • Electrophoresis, Polyacrylamide Gel
  • Histone Acetyltransferases
  • Kinetics
  • Macromolecular Substances
  • Molecular Weight
  • Saccharomyces cerevisiae Proteins*
  • Tetrahymena thermophila / enzymology*

Substances

  • Chromatin
  • Macromolecular Substances
  • Saccharomyces cerevisiae Proteins
  • Durapatite
  • Acetyltransferases
  • Histone Acetyltransferases