Molecular analysis of the interaction between cardosin A and phospholipase Dα

• Autor(a) principal: Simões, Isaura
• Data de Publicação: 2005
• Outros Autores: Mueller, Eva-Christina, Otto, Albrecht, Bur, Daniel, Cheung, Alice Y., Faro, Carlos, Pires, Euclides
• Tipo de documento: Artigo
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10316/8121; https://doi.org/10.1111/j.1742-4658.2005.04967.x

Cardosin A is an RGD-containing aspartic proteinase from the stigmatic papillae of Cynara cardunculus L. A putative cardosin A-binding protein has previously been isolated from pollen suggesting its potential involvement in pollen2013pistil interaction [Faro C, Ramalho-Santos M, Vieira M, Mendes A, Simões I, Andrade R, Verissimo P, Lin X, Tang J & Pires E (1999) J Biol Chem274, 28724201328729]. Here we report the identification of phospholipase D03B1 as a cardosin A-binding protein. The interaction was confirmed by coimmunoprecipitation studies and pull-down assays. To investigate the structural and molecular determinants involved in the interaction, pull-down assays with cardosin A and various glutathione S-transferase-fused phospholipase D03B1 constructs were performed. Results revealed that the C2 domain of phospholipase D03B1 contains the cardosin A-binding activity. Further assays with mutated recombinant forms of cardosin A showed that the RGD motif as well as the unprecedented KGE motif, which is structurally and charge-wise very similar to RGD, are indispensable for the interaction. Taken together our results indicate that the C2 domain of plant phospholipase D03B1 can act as a cardosin A-binding domain and suggest that plant C2 domains may have an additional role as RGD/KGE-recognition domains.