NucleosomeClass3

NucleosomeClass3 - Nucleosomes: what, why and where? Click...

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Click to edit Master subtitle style 3/30/10 Nucleosomes: what, why and where? Rob Brewster
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3/30/10 Outline What is a nucleosome? - how is DNA packaged/organized in Eukaryotes? Why do nucleosomes form? - DNA is stiff, how do ~100 bp loops form so readily? Where do nucleosomes form? - What controls the spacing and structure of nucleosomes on the chromosome? Accessibility of DNA in the nucleosomes - How is DNA inside a nucleosomes accessed?
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3/30/10 DNA Organization in Eukaryotes DNA is packaged and condensed into chromosome - Human genome is big, nucleus is small ~ 2 billion basepairs ≈ 2m nucleus radius ~ 6 µm - Many different levels of organization - Compacts chromosome - Regulates transcription by making portions of the chromosome more/less accessible (up to 80% is inaccessible to protein binding) - We will focus on nucleosome formation (Alberts, Essential Cell (Lee W et al., Nat. Genetics 2007)
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3/30/10 Electron micrograph of chromatin at low ionic strength - Nucleosomes appear as “beads on a string Basic repeating structure can be probed (protect and seq method) - Digestion enzyme cuts accessible regions of DNA - DNA protected by nucleosome is not cut What is a nucleosome? (Electron Micrograph from Olins and Olins) Mon o Du al Mon o Mon o
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3/30/10 Enzymatic digests Particular enzymes can cut double stranded DNA between basepairs - Most have specific recognition sites - Micrococcal nuclease cleaves everything it can (no specific seq.) Dilution of nuclease 1:3 (pictures from
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3/30/10 (Peter J. Russell, iGenetics ) Repeating structure contains 5 different proteins - Main body is an octomer formed from: two copies each of H2A, H2B, H3, H4 - DNA wraps 147 bp (1.75 turns) around the core -14 non specific adhesive contacts with histone (major groove – histone) - H1 attaches to the linker region and changes the conformation; required for chromatin formation Structure of individual nucleosome
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3/30/10 DNA rigidity DNA is stiff… - ~150bp persistence length, λ, for lysed bacterial genome - loops smaller than λ should be rare … but not that stiff - single turn around histone core ~100 bp
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Back of the powerpoint calculation Energy paid to bend loop of 147 bp DNA in 16.5 bp circle Assume λ=150 bp then:
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NucleosomeClass3 - Nucleosomes: what, why and where? Click...

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