Publicação
Understanding the detailed mechanism of interaction between La-related proteins and their RNA targets
| Resumo: | The importance of maturation and processing of all RNA molecules is directly related to RNA-binding proteins (RBPs) and their RNA-binding domains (RBDs). Amongst this class of proteins is the La-related proteins (LaRPs) superfamily, that is characterized by a conserved RNA-binding unit labeled the La-module, which is composed by the La motif (LaM) and the RNA-recognition motif 1 (RRM1). The work presented in this report concerns LaRP4B and its relationship with LaRP4A, since both proteins have identified functions related to cancer and further knowledge about them is crucial in order to find possible treatments. The expression and purification of LaRP4B La-module and RRM1 constructs was successful and allowed to pro-duce 15N RRM for future NMR experiments. While using data from previous experiences, the backbone assignment of the RRM was done with a completion of almost 38% of all residues. This assignment also permitted to identify shifts that belonged to the tag present in the RRM. These findings helped to establish a new purification protocol for the RRM so that the tag can be cleaved by introducing an on-column cleavage step. Finally, by utilizing the chemical shifts of the LaRP4B RRM and La motif, obtained by other group members, a Chemical Shift Index study allowed the comparison between the predicted secondary structure of these domains and the ones from LaRP4A. This comparison showed no major differences between both structures that could indicate why the RNA targets are so different between these two closely related members of the LaRP4 family. Further studies are necessary to unveil the mechanism of inter-action between LaRP4B and RNA. |
|---|---|
| Autores principais: | Colaço, Cláudio |
| Assunto: | La Related Proteins La-module La motif RNA Recognition Motif LaRP4B RNA Binding Proteins |
| Ano: | 2022 |
| País: | Portugal |
| Tipo de documento: | dissertação de mestrado |
| Tipo de acesso: | acesso aberto |
| Instituição associada: | Universidade Nova de Lisboa |
| Idioma: | inglês |
| Origem: | Repositório Institucional da UNL |
| Resumo: | The importance of maturation and processing of all RNA molecules is directly related to RNA-binding proteins (RBPs) and their RNA-binding domains (RBDs). Amongst this class of proteins is the La-related proteins (LaRPs) superfamily, that is characterized by a conserved RNA-binding unit labeled the La-module, which is composed by the La motif (LaM) and the RNA-recognition motif 1 (RRM1). The work presented in this report concerns LaRP4B and its relationship with LaRP4A, since both proteins have identified functions related to cancer and further knowledge about them is crucial in order to find possible treatments. The expression and purification of LaRP4B La-module and RRM1 constructs was successful and allowed to pro-duce 15N RRM for future NMR experiments. While using data from previous experiences, the backbone assignment of the RRM was done with a completion of almost 38% of all residues. This assignment also permitted to identify shifts that belonged to the tag present in the RRM. These findings helped to establish a new purification protocol for the RRM so that the tag can be cleaved by introducing an on-column cleavage step. Finally, by utilizing the chemical shifts of the LaRP4B RRM and La motif, obtained by other group members, a Chemical Shift Index study allowed the comparison between the predicted secondary structure of these domains and the ones from LaRP4A. This comparison showed no major differences between both structures that could indicate why the RNA targets are so different between these two closely related members of the LaRP4 family. Further studies are necessary to unveil the mechanism of inter-action between LaRP4B and RNA. |
|---|