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The binding sites for large and small high-mobility-group (HMG) proteins. Studies on HMG-nucleosome interactions in
Abstract:
Studies in vitro of binding high-mobility-group (HMG) proteins to nucleosomal particles that differ in their DNA contents reflect several aspects pertinent to their function in vivo. Two molecules of HMG 14 or 17 are accommodated by particles with 140 or 180 base pairs of DNA whereas HMG 1 or 2 are only bound by the larger specimens irrespective of the presence of HMG 14/17. It is concluded that one molecule of HMG 1 or 2 binds to the 40 base pairs of linker DNA whereas the HMG 14 or 17 molecules associate with the nucleosomal core. At physiological ionic strength, HMG 14 binding is cooperative, probably by triggering a conformational change in the nucleosomal particle. The phenomenon has been studied by two independent techniques. Besides the common gel-electrophoretic system, a centrifugation assay is described, which permits the derivation of a Hill coefficient nH = 1.3 and dissociation constants in the range of 30-90 nM at 0.15 M NaCl, pH 6.8.
Insights
High-mobility-group (HMG) proteins bind differently to nucleosomes based on DNA content. HMG 1/2 bind linker DNA, while HMG 14/17 bind the nucleosomal core, with HMG 14 showing cooperative binding.
Area of Science:
- Molecular Biology
- Chromatin Structure
- Protein-DNA Interactions
Background:
- High-mobility-group (HMG) proteins are crucial for DNA packaging and gene regulation.
- Nucleosomes, the basic units of chromatin, consist of DNA wrapped around histone proteins.
Purpose of the Study:
- To investigate the binding characteristics of different HMG proteins to nucleosomal particles with varying DNA content.
- To elucidate the specific binding sites and cooperative interactions of HMG proteins within nucleosomes.
Main Methods:
- In vitro studies using nucleosomal particles with defined DNA lengths (140-180 base pairs).
- Gel-electrophoretic and centrifugation assays to analyze protein binding.
- Determination of dissociation constants and Hill coefficients.
Main Results:
- Two molecules of HMG 14 or HMG 17 bind to nucleosomes with 140-180 base pairs of DNA.
- HMG 1 or HMG 2 bind to larger nucleosomal particles (180 base pairs), associating with linker DNA.
- HMG 14/17 associate with the nucleosomal core, and HMG 14 binding is cooperative, suggesting conformational changes.
Conclusions:
- HMG 1/2 proteins bind to the 40 base pairs of linker DNA.
- HMG 14/17 proteins associate with the nucleosomal core.
- HMG 14 exhibits cooperative binding to nucleosomes, indicating a role in modulating chromatin structure.