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dc.contributor.advisorMoore, Stanleyen_US
dc.creatorKlemmer, Kent Conraden_US
dc.date.accessioned2010-10-13T12:13:38Zen_US
dc.date.accessioned2013-01-04T05:01:16Z
dc.date.available2011-11-25T08:00:00Zen_US
dc.date.available2013-01-04T05:01:16Z
dc.date.created2010-10en_US
dc.date.issued2010-10en_US
dc.date.submittedOctober 2010en_US
dc.identifier.urihttp://hdl.handle.net/10388/etd-10132010-121338en_US
dc.description.abstractDosage compensation is the key regulatory process employed in Drosophila melanogaster to equalize the level of gene transcripts between the single X chromosome in males (XY) and the two X chromosomes in females (XX). Dimorphic sex chromosomes evolved by the severe degeneration of the Y chromosome, giving rise to an imbalance between the heterogametic sex and the homogametic sex. Vital to the viability of male Drosophila is the dosage compensation complex (DCC), a ribonucleoprotein complex that mediates the precise two-fold transcription of the single male X chromosome. The DCC is comprised of five proteins: male-specific-lethal proteins (MSL) 1, 2, and 3, male absent-on-the-first (MOF), maleless (MLE), and two non-coding RNAs. The complex specifically co-localizes along the male X chromosome in a reproducible manner, resulting in acetylation of lysine 16 of the N-terminal tail of histone H4. The exact mechanism of recruitment and spreading of the DCC along the male X chromosome remains unclear; recent studies propose a multi-step mechanism involving DNA sequence elements, epigenetic marks, and transcription. Understanding how dosage compensation functions provides insight into the interplay between gene regulation and chromatin remodelling. The goal of this project was to better understand how Drosophila MSL1, MSL3, and MOF interact and how their interaction modulates MOF’s acetyltransferase activity. Recombinant protein constructs were cloned and over-expressed in a bacterial expression system permitting future structure determination by X-ray crystallography. The dMSL1820-1039 construct consisted of the C-terminal domain, reported to be able to interact with both dMSL3 and dMOF. dMSL3186-512 contained the domain required for the interaction with dMSL1 and dMOF. dMOF371-827 was comprised of the catalytic domain, the CCHC zinc finger, and the chromodomain, as the N-terminal region does not encode any known domains. All three recombinant proteins were successfully cloned, over-expressed, and purified to homogeneity. Recombinant dMOF371-827 was determined to acetylate histones. Interaction studies using GST pull-down assays and size exclusion chromatography determined that dMSL1820-1039 and dMOF371-827 did not interact above background levels. Moreover, size exclusion chromatography revealed dMSL3186-512 and dMOF371-827 did not interact nor did the three recombinant proteins form a stable complex.en_US
dc.language.isoen_USen_US
dc.subjectdosage compensationen_US
dc.subjectmale specific lethalen_US
dc.subjectGST-fusion proteinsen_US
dc.titleBiochemical and structural studies of dosage compensation members : MSL1, MSL3, and MOF from Drosophila melanogasteren_US
thesis.degree.departmentBiochemistryen_US
thesis.degree.disciplineBiochemistryen_US
thesis.degree.grantorUniversity of Saskatchewanen_US
thesis.degree.levelMastersen_US
thesis.degree.nameMaster of Science (M.Sc.)en_US
dc.type.materialtexten_US
dc.type.genreThesisen_US
dc.contributor.committeeMemberKhandelwal, Ramjien_US
dc.contributor.committeeMemberBull, Harolden_US
dc.contributor.committeeMemberLee, Jeremyen_US
dc.contributor.committeeMemberBonham, Keithen_US


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