Journal: PLoS ONE
Article Title: Histone H1 Differentially Inhibits DNA Bending by Reduced and Oxidized HMGB1 Protein
Figure Lengend Snippet: The effect of oxidization and mutation of Cys22/Cys44 or Phe37 of HMGB1ΔC on DNA bending. A , the 5´-end 32 P-labeled 123-bp DNA fragment (~1 nM) was pre-incubated with 2, 5, 10, 15, 25, 50 and 100 nM of HMGB1 lacking the acidic C-tail (HMGB1ΔC, left to right ), followed by ligation by T4 DNA ligase (DNA circularization assay). Deproteinised DNA samples were separated by electrophoresis on 5% non-denaturing polyacrylamide gels in 0.5x TBE buffer. B , percentage of DNA circles formed by reduced (black triangle) or oxidized (empty triangle) HMGB1ΔC, as compared to DNA circles formed under the same conditions by reduced (black circles) or oxidized (empty circles) full-length HMGB1. The percentage of the minicircles formed at 100 nM HMGB1 was arbitrary set to 100% (each of the curves represent an average of three independent experiments). C , representative circularization assay using reduced HMGB1ΔC, oxidized HMGB1ΔC, and HMGB1ΔC(F37A). Concentrations of proteins were 5, 10, 25, 50 and 100 nM ( left to right ).
Article Snippet: In agreement with previous reports [ , ], histone H1 could stimulate formation of linear multimers by T4 DNA ligase at low H1-to-DNA ratios.
Techniques: Mutagenesis, Labeling, Incubation, Ligation, Electrophoresis