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dc.contributor.authorBadran, Ahmed Husseinen_US
dc.creatorBadran, Ahmed Husseinen_US
dc.date.accessioned2011-10-25T17:04:57Z
dc.date.available2011-10-25T17:04:57Z
dc.date.issued2010-05
dc.identifier.urihttp://hdl.handle.net/10150/146847
dc.description.abstractCys2-His2 zinc fingers constitute one of the largest classes of DNA-binding domains in the human genome. The modularity of these domains has been recently exploited to design artificial zinc fingers, capable of targeting virtually any sequence. However, the resultant zinc fingers have had significantly high failure rate, owing to low binding affinity and selectivity. Despite much research on the topic, a proper understanding of all the factors involved in zinc finger selectivity, be they natural or artificial, has proved elusive. Here, we present a modification of our previously reported SEquence-Enabled Reassembly (SEER) methodology, allowing us to study zinc finger selectivity with base pair resolution. Using this modified strategy, we show that the natural 3-finger zinc finger Zif268 binding to its consensus site is highly dependent on the availability of specific base pairs, or 'hot spots,' and independent of mutations at adjacent positions. Additionally, we show that positional interdependence plays a large role in the selectivity of both Zif268 and the artificial 6-finger zinc finger Aart for their respective targets. We envision that this technique can be easily applied towards the interrogation of any DNA-binding domain in a high throughput and accurate manner.
dc.language.isoenen_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.titleSplit-Protein Systems for the Detection and Interrogation of Protein-Nucleic Acid Interactionsen_US
dc.typetexten_US
dc.typeElectronic Thesisen_US
thesis.degree.grantorUniversity of Arizonaen_US
thesis.degree.levelbachelorsen_US
thesis.degree.disciplineHonors Collegeen_US
thesis.degree.disciplineBiochemistry and Molecular Biophysicsen_US
thesis.degree.nameB.S.en_US
refterms.dateFOA2018-05-27T22:33:50Z
html.description.abstractCys2-His2 zinc fingers constitute one of the largest classes of DNA-binding domains in the human genome. The modularity of these domains has been recently exploited to design artificial zinc fingers, capable of targeting virtually any sequence. However, the resultant zinc fingers have had significantly high failure rate, owing to low binding affinity and selectivity. Despite much research on the topic, a proper understanding of all the factors involved in zinc finger selectivity, be they natural or artificial, has proved elusive. Here, we present a modification of our previously reported SEquence-Enabled Reassembly (SEER) methodology, allowing us to study zinc finger selectivity with base pair resolution. Using this modified strategy, we show that the natural 3-finger zinc finger Zif268 binding to its consensus site is highly dependent on the availability of specific base pairs, or 'hot spots,' and independent of mutations at adjacent positions. Additionally, we show that positional interdependence plays a large role in the selectivity of both Zif268 and the artificial 6-finger zinc finger Aart for their respective targets. We envision that this technique can be easily applied towards the interrogation of any DNA-binding domain in a high throughput and accurate manner.


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