H-DNA and d(TC)n binding proteins

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dc.contributor.advisorVan de Sande, Johan H.
dc.contributor.authorYee, Harris A.
dc.date.accessioned2005-07-27T23:32:05Z
dc.date.available2005-07-27T23:32:05Z
dc.date.issued1991
dc.descriptionBibliography: p. 133-143.en
dc.description.abstractRepetitive d(TC)n tracts have been shown in vitro to adopt an unusual DNA conformation, H-DNA. H-DNA is an intramolecular triplex favored to form in sequences with mirror symmetry and having either a homopurine and /or homopyrmidine strand bias. These d(TC)n tracts are prevalently found in eukaryotic genomes. Specifically, these sequences have been isolated in the regulatory regions of genes, sites of recombination and origins of replication. Many biological roles have been proposed and a putative role in transcription has been demonstrated. The adventitious multi-functional aspect of this sequence strongly suggests a more global role in the structure and function relationship, such as in the organization of chromatin and the nuclear matrix. If these sequences have biological functions in the cell, the question is whether these functions include the formation of triple stranded structures. Two d(TC)n·d(GA)n bearing plasmids, pTClS and pTC32 have been constructed to examine the relationships between superhelical density, protonation and insert size in the context of H-DNA formation. The former plasmid contains a synthetic d(TC)is insert and the latter plasmid contains a synthetic d(TCh2 insert. Two dimensional topoisomer electrophoretic analysis is a useful and powerful techique for examining thermodynamically stable transitions. From these results, thermodynamic parameters were calculated. The formation of H-DNA for d(TC)n sequences is favored under acidic conditions and is length dependent. The pattern of Sl nuclease activity for pTC15 was consistent with H-DNA and for pTC32 was not consistent with a single or predominant conformer of H-DNA but a mixed population of conformers. A natural sequence with a polypyrmidine strand bias derived from mouse was characterized to form H-DNA in vitro. The cloned insert contains a (CT)io insert flanked by (T ,C) rich tracts. The conditions for H-DNA formation of this mouse-derived sequence is similar to the conditions of the synthetic insert cloned within pTC15. A protein-DNA interaction with the sequence d(TC)0 was examined using nuclear protein extracts. A cluster of proteins was discovered to bind to the single­stranded oligonucleotide d(TC)i6 but not its complementary strand, d(GA)l6 nor the duplex sequence d(TCh6·d(GAh6- These single-stranded d(TC)n binding proteins are found among several mammalian species examined and have been sized as 55.5 and 57 kD doublets using southwestern analysis. The existence of this protein is incompatible with postulated in vivo existence of the d(TC)n type triplexes in eukaryotic cells.
dc.format.extentxx, 143 leaves ; 30 cm.en
dc.identifier.citationYee, H. A. (1991). H-DNA and d(TC)n binding proteins (Master's thesis, University of Calgary, Calgary, Canada). Retrieved from https://prism.ucalgary.ca. doi:10.11575/PRISM/19749en_US
dc.identifier.doihttp://dx.doi.org/10.11575/PRISM/19749
dc.identifier.isbn0315669233en
dc.identifier.lccQP 551 Y43 1991en
dc.identifier.urihttp://hdl.handle.net/1880/24513
dc.language.isoeng
dc.publisher.institutionUniversity of Calgaryen
dc.publisher.placeCalgaryen
dc.rightsUniversity of Calgary graduate students retain copyright ownership and moral rights for their thesis. You may use this material in any way that is permitted by the Copyright Act or through licensing that has been assigned to the document. For uses that are not allowable under copyright legislation or licensing, you are required to seek permission.
dc.subject.lccQP 551 Y43 1991en
dc.subject.lcshProtein binding
dc.subject.lcshProteins - Affinity labeling
dc.titleH-DNA and d(TC)n binding proteins
dc.typemaster thesis
thesis.degree.disciplineUniversity Biochemistry Group
thesis.degree.grantorUniversity of Calgary
thesis.degree.nameMaster of Science (MSc)
ucalgary.item.requestcopytrue
ucalgary.thesis.accessionTheses Collection 58.002:Box 806 520535239
ucalgary.thesis.notesoffsiteen
ucalgary.thesis.uarcreleaseyen
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