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dc.contributor.advisorHausrath, Andrew
dc.contributor.authorMumme, Sherilyn Nicoleen_US
dc.creatorMumme, Sherilyn Nicoleen_US
dc.date.accessioned2013-08-09T16:53:50Z
dc.date.available2013-08-09T16:53:50Z
dc.date.issued2013
dc.identifier.urihttp://hdl.handle.net/10150/297722
dc.description.abstractThe mumps Nucleocapsid-Binding Domain (NBD) protein is a small protein that mediates attachment of the viral polymerase to the nucleocapsid. Mumps NBD lacks tertiary structure in isolation; however, it folds into a specific three-dimensional structure when interacting with the N protein. The lack of structure is thought to weaken this binding, facilitating forward motion of the polymerase. Since the closely related NBD from measles is extremely stable without binding, we hypothesize that the mumps NBD may have accumulated mutations that reduce its stability. The mumps NBD is an attractive protein to study the determinants of stability due to the latent capacity to be stabilized. One approach to investigating stability determinants is to create multiple variants, and screen for those which have decreased proteolytic susceptibility indicating increased stability. In this work, we investigated the applicability of pulse proteolysis to screen variants of the mumps NBD. The mumps NBD was mutated using Error Prone PCR. Three mutated mumps NBD variants were then compared to the wild-type in order to determine information about the relative proteolytic susceptibility of the mutants. The use of different concentrations of urea during the proteolysis was investigated as a means to increase the observed differences between variants.
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.titleUse of Pulse Proteolysis to Investigate the Stability Determinants of the Mumps Nucleocapsid-Binding Domainen_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.disciplineBiochemistryen_US
thesis.degree.nameB.S.en_US
refterms.dateFOA2018-08-30T09:52:52Z
html.description.abstractThe mumps Nucleocapsid-Binding Domain (NBD) protein is a small protein that mediates attachment of the viral polymerase to the nucleocapsid. Mumps NBD lacks tertiary structure in isolation; however, it folds into a specific three-dimensional structure when interacting with the N protein. The lack of structure is thought to weaken this binding, facilitating forward motion of the polymerase. Since the closely related NBD from measles is extremely stable without binding, we hypothesize that the mumps NBD may have accumulated mutations that reduce its stability. The mumps NBD is an attractive protein to study the determinants of stability due to the latent capacity to be stabilized. One approach to investigating stability determinants is to create multiple variants, and screen for those which have decreased proteolytic susceptibility indicating increased stability. In this work, we investigated the applicability of pulse proteolysis to screen variants of the mumps NBD. The mumps NBD was mutated using Error Prone PCR. Three mutated mumps NBD variants were then compared to the wild-type in order to determine information about the relative proteolytic susceptibility of the mutants. The use of different concentrations of urea during the proteolysis was investigated as a means to increase the observed differences between variants.


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