The involvement of histone acetylation in facilitating gene expression is well-established,
particularly in the case of histones H3 and H4. It was previously shown in Saccharomyces
cerevisiae that gene expression was significantly down-regulated and chromatin more
condensed in stationary phase compared to exponential phase. We were therefore
interested in establishing the acetylation state of histone H3 and H4 in stationary and in
exponential phase, since the regulation of this modification could contribute to transcriptional
shut-down and chromatin compaction during semi-quiescence. We made use of nano-spray
tandem mass spectrometry to perform a precursor ion scan to detect an m/z 126 immonium
ion, diagnostic of an N -acetylated lysine residue that allowed unambiguous identification of
acetylated as opposed to tri-methylated lysine. The fragmentation spectra of peptides thus
identified were searched with Mascot against the Swiss-Prot database, and the y-ion and bion
fragmentation series subsequently analyzed for mass shifts compatible with acetylated
lysine residues. We found that K9, K14 and K36 of histone H3 and K12 and K16 of histone
H4 were acetylated in exponential phase (bulk histones), but could not detect these
modifications in histones isolated from stationary phase cells. The corresponding unacetylated
peptides were, however, observed. This result was confirmed by Western
analysis (work not presented here). H4K16 acetylation was previously shown to disrupt
formation of condensed chromatin in vitro.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:ufs/oai:etd.uovs.ac.za:etd-10192011-140304 |
| Date | 19 October 2011 |
| Creators | Ngubo, Mzwanele |
| Contributors | Prof H Patterton |
| Publisher | University of the Free State |
| Source Sets | South African National ETD Portal |
| Language | en-uk |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.uovs.ac.za//theses/available/etd-10192011-140304/restricted/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to University Free State or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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