Proteasome activator subunit PA28α and related Ki antigen (PA28γ) are absent from the nuclear fraction purified by sucrose gradient centrifugation

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Abstract

The aim of the present work was to attempt to partially purify PA28 (REG) α and γ (Ki antigen) in the nuclear fraction from NT2/D1 cells. Nuclei were isolated by the hypertonic sucrose gradient centrifugation method and fractionated into membrane/nucleoplasmic and chromatin/nucleolar fractions. Western blotting with anti-histone and anti-β-tubulin monoclonal antibodies confirmed the accuracy of the procedure. Proteasomes were present mainly in the cytoplasm but also in the nuclei. Disruption of the nuclear envelope released the proteasomes implying a loose or no binding with the chromatin. PA28α and γ were detected mainly in the cytosol and to a lesser extent in the crude nuclear pellet, however the purified nuclei were devoid of PA28α and γ. This indicates, that only a small fraction of the PA28 activator is present in the nuclei as detected by immunofluorescence or/and it is easily removed during nuclear purification.

Original languageEnglish (US)
Pages (from-to)273-276
Number of pages4
JournalInternational Journal of Biochemistry and Cell Biology
Volume31
Issue number2
DOIs
Publication statusPublished - Feb 1999
Externally publishedYes

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Keywords

  • NT2 cells
  • Nucleus
  • PA28
  • Proteasome
  • Proteasome activator

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology

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