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Investigations into the role of histone H2A ubiquitination in chromatin
Bibliography: leaves 141-150.
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| Format: | Thesis |
| Language: | English |
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Department of Molecular and Cell Biology
2014
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| _version_ | 1867613341691674624 |
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| access_status_str | Open Access |
| author | Jason, Laure Jeanine Monique |
| author2 | Lindsey, George G |
| author_browse | Jason, Laure Jeanine Monique Lindsey, George G |
| author_facet | Lindsey, George G Jason, Laure Jeanine Monique |
| author_sort | Jason, Laure Jeanine Monique |
| collection | Thesis |
| description | Bibliography: leaves 141-150. |
| format | Thesis |
| id | oai:open.uct.ac.za:11427/9765 |
| institution | University of Cape Town (South Africa) |
| language | eng |
| last_indexed | 2026-06-10T12:34:36.552Z |
| license_str | Not specified — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from UCTD — University of Cape Town Open Access Repository |
| publishDate | 2014 |
| publishDateRange | 2014 |
| publishDateSort | 2014 |
| publisher | Department of Molecular and Cell Biology |
| publisherStr | Department of Molecular and Cell Biology |
| record_format | dspace |
| source_str | UCTD — University of Cape Town Open Access Repository |
| spelling | oai:open.uct.ac.za:11427/9765 Investigations into the role of histone H2A ubiquitination in chromatin Jason, Laure Jeanine Monique Lindsey, George G Brandt, Wolf F Patterton, Hugh Biochemistry Bibliography: leaves 141-150. An in vitro system was used to determine the effect of histone H2A ubiquitination on linker histone binding to mononucleosomes. Hybrid octamers containing either H2A or ubiquitinated H2A (uH2A) were reconstituted onto random sequence 167 bp DNA. The affinity of the resultant nucleosome cores for linker histone H1 was determined from nucleoprotein gel shifts, protein analyses and thermal denaturation. Ubiquitinated H2A did not inhibit linker histone binding to nucleosome cores. The effect of uH2A on nucleosome and chromatosoine positioning on a 208 bp Lytechinus variegatus 5S rDNA fragment was investigated using a combination of micrococcal nuclease digestion and subsequent restriction enzyme digestion of the core particle or chromatosome DNA. Nucleosomes and chromatosomes containing uH2A were found to occupy the same positions on the template DNA as those containing H2A. Chromatin folding of nucleosomal arrays containing either H2A or uH2A was analysed using a quantitative agarose gel electrophoresis system developed by Hansen and co-workers. The extent of folding of nucleosomal arrays containing uH2A was comparable to that of control nucleosomal arrays. A differential centrifugation assay was used to monitor the extent of divalent cation induced oligomerisation of reconstituted nucleosomal arrays. Nucleosomal arrays containing uH2A were found to oligomerise at a lower magnesium concentration than control arrays. As a first step towards studying the effects of H2A ubiquitination in linker histone-bound nucleosomal arrays, a novel method for linker histone reconstitution onto long chromatin stripped of linker histones was developed. The fidelity of linker histone reconstitution was assayed by micrococcal nuclease digestion, thermal denaturation and determination of the orientation of neighbouring linker histone molecules in extended chromatin. In a separate study, the relationship between the observed repeat length of chromatin and the rate of micrococcal nuclease digestion was investigated. The repeat length of the same starting chromatin preparation at equivalent extents of digestion was found to vary according to the rate of digestion. 2014-11-21T16:09:50Z 2014-11-21T16:09:50Z 1999 Doctoral Thesis Doctoral PhD http://hdl.handle.net/11427/9765 eng application/pdf Department of Molecular and Cell Biology Faculty of Science University of Cape Town |
| spellingShingle | Biochemistry Jason, Laure Jeanine Monique Investigations into the role of histone H2A ubiquitination in chromatin |
| thesis_degree_str | Doctoral |
| title | Investigations into the role of histone H2A ubiquitination in chromatin |
| title_full | Investigations into the role of histone H2A ubiquitination in chromatin |
| title_fullStr | Investigations into the role of histone H2A ubiquitination in chromatin |
| title_full_unstemmed | Investigations into the role of histone H2A ubiquitination in chromatin |
| title_short | Investigations into the role of histone H2A ubiquitination in chromatin |
| title_sort | investigations into the role of histone h2a ubiquitination in chromatin |
| topic | Biochemistry |
| url | http://hdl.handle.net/11427/9765 |
| work_keys_str_mv | AT jasonlaurejeaninemonique investigationsintotheroleofhistoneh2aubiquitinationinchromatin |