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dc.contributor.authorNadkarni, Ashwini
dc.date.accessioned2014-03-04T02:28:30Z
dc.date.available2014-03-04T02:28:30Z
dc.date.issued2006-12
dc.identifier.othernadkarni_ashwini_u_200612_phd
dc.identifier.urihttp://purl.galileo.usg.edu/uga_etd/nadkarni_ashwini_u_200612_phd
dc.identifier.urihttp://hdl.handle.net/10724/23697
dc.description.abstractThe empirical process of macromolecular crystallization has always been a bottleneck in the production of protein structures. Co-crystallization of intransigent proteins with antibody fragments has recently emerged as a powerful tool to facilitate crystal formation and improve crystal quality. However, this approach is not readily applied because of the lack of adequate quantities of antibody fragments that recognize target proteins and also due to the complexity of the selection system. The objective of this dissertation was to develop a recombinant antibody fragment (Fab)-based system for the high-throughput generation of co-crystallization proteins (CCPs). Phage display technology lies at the heart of the selection process, but the method has been notoriously slow and tedious. A rapid approach was developed that was faster than standard methods and because of the simplified manipulations, less tedious and easily applicable to robotic automation. The other focus of this dissertation was on addressing the underlying issues of producing antibody fragments in a bacterial system. Automated methods of purifying the target-specific Fabs were developed that are general in nature and should work with all of the Fabs produced in this system. By optimizing the bacterial strain, culture conditions, and purification strategy, as much as 12 mg of highly purified Fab was produced per liter of bacterial cell culture in less than two days using bench-top equipment.
dc.languageeng
dc.publisheruga
dc.rightspublic
dc.subjectAntibody Fragment
dc.subjectCo-crystallization
dc.subjectCrystallization
dc.subjectFab
dc.subjectHigh-throughput
dc.subjectPhage Display
dc.subjectSecretion
dc.subjectCytoplasmic
dc.subjectStructural Genomics
dc.titleDevelopment of high-throughput approaches for the phage display generation and production of recombinant antibody fragments for macromolecular co-crystallization
dc.typeDissertation
dc.description.degreePhD
dc.description.departmentPharmaceutical and Biomedical Sciences
dc.description.majorPharmacy
dc.description.advisorCory Momany
dc.description.committeeCory Momany
dc.description.committeeRoger G. Dean
dc.description.committeeArthur Grider
dc.description.committeeJ. Warren Beach
dc.description.committeeShelley Hooks


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