Linker histone partial phosphorylation: effects on secondary structure and chromatin condensation.
Bottom Line: Infrared spectroscopy analysis showed a gradual increase of β-structure in the phosphorylated samples, concomitant to a decrease in α-helix/turns, with increasing linker histone phosphorylation.A decrease of the sedimentation rate through sucrose gradients of the phosphorylated samples was observed, indicating a global relaxation of the 30-nm fiber following linker histone phosphorylation.Phosphorylated chromatin had lower percentages in volume of aggregated molecules and the aggregates had smaller hydrodynamic diameter than unphosphorylated chromatin, indicating that linker histone phosphorylation impaired chromatin aggregation.
Affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Biociencias, Universidad Autónoma de Barcelona, 08193, Bellaterra, Barcelona, Spain.Show MeSH
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Mentions: In order to observe the effects of phosphorylation on chromatin aggregation, samples were incubated in the presence of CDK2 for 1 h, 5 h and overnight at 30°C. Incubation at 30°C induced, by itself, the aggregation of part of the sample in a time-dependent manner when chromatin was in the presence of 1.6-mM MgCl2 (Supplementary Figure S8C). However, no significant aggregation was observed in 1-mM MgCl2 upon incubation at 30°C (Supplementary Figure S8B). Chromatin aggregation was thus dependent at the same time on the presence of a critical concentration of Mg2+ ions and the temperature and time of incubation. The effects of phosphorylation on chromatin aggregation were, therefore, evaluated in comparison with the degree of aggregation of samples incubated during the same time periods at 30°C, but without kinase (Figures 5 and 6 and Supplementary Table S6).
Affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Biociencias, Universidad Autónoma de Barcelona, 08193, Bellaterra, Barcelona, Spain.