Action of recombinant human apoptotic endonuclease G on naked DNA and chromatin substrates: cooperation with exonuclease and DNase I.
Endonuclease G (endoG) is released from mitochondria during apoptosis and is in part responsible for internucleosomal DNA cleavage. Here we report the action of the purified human recombinant form of this endonuclease on naked DNA and chromatin substrates. The addition of the protein to isolated nuclei from non-apoptotic cells first induces higher order chromatin cleavage into DNA fragments > or = 50 kb in length, followed by inter- and intranucleosomal DNA cleavages with products possessing significant internal single-stranded nicks spaced at nucleosomal ( approximately 190 bases) and subnucleosomal ( approximately 10 bases) periodicities. We demonstrate that both exonucleases and DNase I stimulate the ability of endoG to generate double-stranded DNA cleavage products at physiological ionic strengths, suggesting that these activities work in concert with endoG in apoptotic cells to ensure efficient DNA breakdown.
Widlak
P
P
Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, Texas 75390, USA.
Li
L Y
LY
Wang
X
X
Garrard
W T
WT
D002384:Catalysis; D002843:Chromatin / Q000378:metabolism*; D004247:DNA / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000378:metabolism*; D004706:Endodeoxyribonucleases / Q000302:isolation & purification / Q000378:metabolism*; D005092:Exonucleases / Q000378:metabolism*; D006801:Humans; D009994:Osmolar Concentration; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D013379:Substrate Specificity
D002843:Chromatin; D011994:Recombinant Proteins; D004247:DNA; D004706:Endodeoxyribonucleases; D005092:Exonucleases; C052391:endonuclease G; D003850:Deoxyribonuclease I