RESUMO
Conserved immune cell signaling in fish was recently highlighted by the identification of various immune cell signaling molecules. Tumor necrosis factor (TNF) receptor-associated factor (TRAF) proteins are critical adaptor molecules in immune cell signaling and contain E3 ubiquitin ligase activity. Here, we report the first crystal structure of the TRAF5 TRAF domain from the black rockcod (Notothenia coriiceps; ncTRAF5). Our structure revealed both similarities and differences with mammalian TRAF5. Structural and biochemical analyses indicated that ncTRAF5 forms a functional trimer unit in solution, with a structural flexibility that might be critical for imparting resistance to cold temperature-induced stress. We also found conserved surface residues on ncTRAF5 that might be critical binding hot spots for interaction with various receptors.
Assuntos
Doenças dos Peixes/imunologia , Imunidade Inata/genética , Perciformes/genética , Perciformes/imunologia , Fator 5 Associado a Receptor de TNF/genética , Fator 5 Associado a Receptor de TNF/imunologia , Sequência de Aminoácidos , Animais , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Alinhamento de Sequência/veterinária , Transdução de Sinais , Fator 5 Associado a Receptor de TNF/químicaRESUMO
Multi-functional transglutaminase 2 (TG2), which possesses protein cross-linking and GTP hydrolysis activities, is involved in various cellular processes, including apoptosis, angiogenesis, wound healing, and neuronal regeneration, and is associated with many human diseases, including inflammatory disease, celiac disease, neurodegenerative disease, diabetes, tissue fibrosis, and cancers. Although most biochemical and cellular studies have been conducted with the TG2 (G224) form, the TG2 (G224V) form has recently emerged as a putative natural variant of TG2. In this study, we characterized the putative natural form of TG2, TG2 (G224V), and through a new crystal structure of TG2 (G224V), we revealed how TG2 (G224V) gained stability and higher Ca2+-dependent activity than an artificial variant of TG2 (G224).
Assuntos
Proteínas de Ligação ao GTP/química , Proteínas de Ligação ao GTP/metabolismo , Transglutaminases/química , Transglutaminases/metabolismo , Substituição de Aminoácidos , Cálcio/metabolismo , Domínio Catalítico/genética , Cristalografia por Raios X , Estabilidade Enzimática/genética , Proteínas de Ligação ao GTP/genética , Variação Genética , Humanos , Cinética , Modelos Moleculares , Mutagênese Sítio-Dirigida , Conformação Proteica , Proteína 2 Glutamina gama-Glutamiltransferase , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Transglutaminases/genéticaRESUMO
The apoptosis repressor with caspase-recruiting domain (ARC) is aberrantly overexpressed in various cancers. ARC contains a caspase recruitment domain (CARD) that is the main mediator of protein-protein interactions. Mutation of Leu31 within the CARD of ARC to Phe (ARC_L31F) is widely used as a functionally defective mutant of ARC despite a lack of clear experimental evidence regarding how its functionality is lost. In this study, we show that L31 in helix 2 (H2) is critical for stabilization of the helix bundle fold in the CARD domain. In addition, the L31F mutation disrupts homodimer formation that is critical to ARC functions. Our current study reveals the molecular basis for the L31F mutation disrupting the ARC CARD functions.