The characterization of the possible interaction between the Chlamydia effector tarp and its outative chaperone CT043

• Autor(a) principal: Pedrosa, António José Santos Tedim Sousa
• Data de Publicação: 2011
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10400.14/9267

Chlamydia trachomatis is the most common cause of sexually transmitted bacterial infection in Europe and in the United States. It is a Gram negative obligate intracellular pathogen with a unique biphasic development cycle, possessing two distinct morphological forms, the elementary body (EB) and the reticulate body (RB). The EBs are the infectious form and are able to attach and invade mammalian cells. This process is likely dependent on the Type III Secretion System, which allows secretion of Chlamydia effectors that modulate the actin skeleton of the host cell and promote invasion. One of the first effectors to be secreted is TarP (Translocated Actin Recruiting Protein). Orthologs of this effector are present in all Chlamydia with some polymorphisms associated with some subdomains. An example is the tyrosine rich domain, which is only present in the TarP homolog of the C. trachomatis species. Despite this difference, all chlamydial TarP are able to recruit and nucleate actin to promote the internalisation of Chlamydia. Since early secretion of TarP is crucial for invasion it was hypothesized that this secretion, in order to be efficient must be facilitated by a Type III Secretion chaperone. These usually are small proteins (13-16 kDa) with an acidic pI and a secondary structure motif of α-β-β-β-α-β-β. Three putative Chlamydia trachomatis Type III secretion chaperones, CT043 (Slc1), CT663 (Slc2), CT088 (Scc1), which are present in EBs have been predicted by their secondary structures. The interaction of these chaperones with the N-terminal domain of TarP (amino acids 1-200) was tested using pull-down assays. A specific interaction between TarP and Slc1 was observed, but not with Slc2 or Scc1. Using a bacterial two-hybrid assay, we evaluated the effects of mutating TarP(1-100) on its interaction with Slc1. The objective was to identify critical amino acid residues involved in chaperone binding. Using several mutants TarP derivatives, we were able to conclude that the interaction between TarP and Slc1 may depend of both specific amino acids and hydrophobic interactions.