The Chd1 chromatin remodeler forms long-lived complexes with nucleosomes in the presence of ADP·BeF3- and transition state analogs.
J Biol Chem
; 294(48): 18181-18191, 2019 11 29.
Article
em En
| MEDLINE
| ID: mdl-31636125
ABSTRACT
Chromatin remodelers use helicase-like ATPase domains to reorganize histone-DNA contacts within the nucleosome. Like other remodelers, the chromodomain helicase DNA-binding protein 1 (Chd1) remodeler repositions nucleosomes by altering DNA topology at its internal binding site on the nucleosome, coupling different degrees of DNA twist and DNA movement to distinct nucleotide-bound states of the ATPase motor. In this work, we used a competition assay to study how variations in the bound nucleotide, Chd1, and the nucleosome substrate affect stability of Chd1-nucleosome complexes. We found that Chd1-nucleosome complexes formed in nucleotide-free or ADP conditions were relatively unstable and dissociated within 30 s, whereas those with the nonhydrolyzable ATP analog AMP-PNP had a mean lifetime of 4.8 ± 0.7 min. Chd1-nucleosome complexes were remarkably stable with ADP·BeF3- and the transition state analogs ADP·AlFX and ADP·MgFX, being resistant to competitor nucleosome over a 24-h period. For the tight ADP·BeF3--stabilized complex, Mg2+ was a critical component that did not freely exchange, and formation of these long-lived complexes had a slow, concentration-dependent step. The ADP·BeF3--stabilized complex did not require the Chd1 DNA-binding domain nor the histone H4 tail and appeared relatively insensitive to sequence differences on either side of the Widom 601 sequence. Interestingly, the complex remained stable in ADP·BeF3- even when nucleosomes contained single-stranded gaps that disrupted most DNA contacts with the guide strand. This finding suggests that binding via the tracking strand alone is sufficient for stabilizing the complex in a hydrolysis-competent state.
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Assunto principal:
Saccharomyces cerevisiae
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DNA Fúngico
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Nucleossomos
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Difosfato de Adenosina
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Proteínas de Saccharomyces cerevisiae
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Proteínas de Ligação a DNA
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Fluoretos
Idioma:
En
Revista:
J Biol Chem
Ano de publicação:
2019
Tipo de documento:
Article