Autor(es):
Espadinha, Daniel António Burgos 
Data: 2009
Identificador Persistente: http://hdl.handle.net/10451/1484
Origem: Repositório da Universidade de Lisboa
Assunto(s): Imunologia molecular; Escherichia coli; Técnicas biológicas; Teses de mestrado
Descrição
Tese de mestrado, Biologia (Biologia Humana e Ambiente), 2009, Universidade de Lisboa, Faculdade de Ciências Activation-induced cytidine deaminase (AID) is responsible for the induction of three reactions of DNA somatic modification employed by jawed vertebrates in the context of adaptive immunity: Somatic Hypermutation (SHM), Class Switch Recombination (CSR) and Immunoglobulin Gene Conversion (Ig GC). However, in conditions of deregulation, AID has also been implicated in both lymphoid and non-lymphoid neoplasias. One of most puzzling questions is how the activity of this enzyme is controlled and, in normal conditions, restricted to act only on the Ig loci. Many structural and functional aspects of AID have been discovered by analyzing mutations in AID-impaired patients that have Hyper-IgM syndrome type II. Nevertheless, this approach seems to be exhausted and over the last few years it has not produced new relevant findings. Here, I employ the exact opposite strategy. Since there are no patients with mutated genes encoding hypermorphic AID forms, probably because these would have severe genotoxic and oncogenic effects, I am designing a directed evolution experiment of AID to isolate mutants with enhanced activity. Thus, I plan to unleash the mutagenic potential of this deaminase by mimicking the process of natural evolution in culture, at the protein level, through the reiteration of two main events: Diversification & Selection. I anticipate that the high-activity mutants will result from a number of different possibilities, including the bypass of regulatory mechanisms acting at the protein level. If so, by studying in detail the isolated mutations it should be possible to infer interactions with AID regulatory proteins. The mutants will also be valid tools to create novel cancer murine models, more heuristic and refined than the previous ones. Finally, the mutants will be relevant for several biotechnological applications. Resumo alargado em português disponível no documento
