The primary objective of this dissertation was to combine the power of bioinformatics with synthetic genomics, reverse genetics, and molecular genetic approaches to generate a platform technology, with which to empirically test well-informed hypotheses towards understanding complex mechanisms of viral pathogenesis and replication. This integrative strategy was used to: (1) identify unique sequence signatures that were associated with aberrant or altered gene function, focusing on replicase and structural proteins of the Coronavirus family and the capsid protein of the Norovirus family; (2) use these sequence signatures to generate hypotheses and predictions about gene function or evolution; and then (3) empirically test these models using reverse genetics, synthetic biology, molecular genetics, and biochemistry in the laboratory setting. Applying this integrative strategy allowed us to generate three informative models. First, we generated an informative model for the pleiotropic role of a Coronavirus non-structural protein 10 in replication and proteolytic processing, by demonstrating that this protein regulates RNA transcription during replication, in addition to an essential role in polyprotein processing. Second, we demonstrated that the receptor-binding domain of the Coronavirus spike protein is the minimal domain requiring adaptation for host range switching, which helps explain the evolutionary epidemiology of SARS-CoV emergence from zoonotic reservoirs. And finally, we generated an informative model for the molecular mechanisms governing the persistence of the GII.4 noroviruses in human populations, whereby we demonstrated that these viruses persist by evolving unique epitopes on the capsid protein surface to circumvent herd immunity and mediate receptor switching.Pond, S. L. K., S. D. W. Frost, and S. V. Muse. 2005. HyPhy: ... NCBI Reference Sequence project: update and current status. Nucleic Acids Res ... EMBOSS: The European Molecular Biology Open Software Suite Made Easy: A How-To Manual , Third Edition. Trends in ... 2002. PROSITE: a documented database using patterns and profiles as motif descriptors. Brief Bioinform 3:265-74. 37. Szabadka , Z.
|Title||:||Computational and Molecular Biology Approaches to Viral Replication and Pathogenesis|
|Author||:||Eric F. Donaldson|
|Publisher||:||ProQuest - 2008|