ACF Remodeling Enzyme Complex Description: Recombinant Drosophila ACF Remodeling Enzyme Complex, assembled from Acf1-FLAG and ISWI produced in SF9 cells. ACF is an ATP-dependent chromatin remodeling complex that regulates nucleosome spacing.
ACF Remodeling Enzyme Complex Formulation: ACF Complex at 0.18 mg/ml in 5.2 μl of 20 mM Tris HCl pH 7.9, 150 mM NaCl, 2 mM MgCl2, 0.2 mM EDTA, 10 mM beta-glycerophosphate, 0.01% NP-40, 0.2 mM PMSF, 0.5 mM benzamidine, 1 mM DTT, 15% glycerol, 2 μg/ml leupeptin, 1 μg/ml aprotinin, 0.4 mg/ml recombinant human insulin, 0.4 mg/ml 3x FLAG peptide. Molarity: 973 nM
Storage and Stability: Stable for six months at -80°C from date of receipt. For best results, aliquot and avoid multiple freeze/thaws.
Application Notes: This product is sufficient to perform 100 remodeling reactions using EpiDyne®-FRET substrate. A single reaction is defined as 10 μl containing [5 nM ACF Complex/ 20 nM EpiDyne®-FRET/ 50 μM rATP] and remodels to completion in <20 minutes. 5x ACF Remodeling Assay Buffer is included (100 mM Tris HCl, pH 7.5, 250 mM KCl, 15 mM MgCl2, 0.05% (w/v) BSA, 0.05% (v/v) Tween 20). For best results, aliquot ACF into single-use aliquots upon arrival. Avoid multiple freeze/thaw cycles. For further information, please contact firstname.lastname@example.org or refer to the EpiDyne®-FRET Technote (TN#106; available at www.epicypher. com/technotes).
Fyodorov DV & Kadonaga JT (2002) Nature 418: 897-900 (PMID 12192415)
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ATP-dependent Chromatin Remodeling Assay: EpiDyne®-FRET Chromatin Remodeling Substrate (20 nM; Catalog No. 16-4201) incubated with ACF Remodeling Enzyme Complex (concentrations indicated), with or without ATP (50 μM). Curves denote FRET efficiency/chromatin remodeling. (Click image to enlarge)
Chromatin Remodeling Assay:Polysomes treated with 2 different MNase concentrations and the indicated amounts of chromatin remodeler: No Remodeler, ACF Complex, or ISWI alone. M indicates 123 bp molecular weight markers. (Click image to enlarge)