RESUMO
The immunomodulatory activity of plant lectins has been evaluated because of their high selectivity for glycans linked to receptors on innate and adaptative immune cells. ArtinM is a mannosyl-binding lectin, obtained from the seeds of Artocarpus heterophyllus, that induces the differentiation of CD4+ T cells and macrophages by interacting with CD3 and TLR2/CD14, respectively. This ArtinM property ultimately favors the combat of intracellular pathogens, opening new perspectives on the lectins application as immunomodulatory agents. The current section describes protocols for purification and evaluation of ArtinM biological activity. The purification is based on the ArtinM-D-mannose affinity. The effect of inducing IL-12 production by murine macrophages cell line is adopted to evaluate the ArtinM biological activity.
Assuntos
Artocarpus/metabolismo , Linfócitos T CD4-Positivos/citologia , Fatores Imunológicos/farmacologia , Macrófagos/citologia , Lectinas de Plantas/farmacologia , Animais , Artocarpus/química , Complexo CD3/metabolismo , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/metabolismo , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Fatores Imunológicos/isolamento & purificação , Interleucina-12/metabolismo , Receptores de Lipopolissacarídeos/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Manose/metabolismo , Camundongos , Lectinas de Plantas/isolamento & purificação , Células RAW 264.7 , Sementes/química , Sementes/metabolismo , Receptor 2 Toll-Like/metabolismoRESUMO
Some lectins of pathogens interact with host cells through the recognition of specific carbohydrates displayed on the mammals' cell surface. The microneme protein 1 (MIC1) from Toxoplasma gondii has a lectin domain that specifically binds sialic acid residues, often found in the terminal positions of N-glycans of mammalian cells. The necessary studies on the MIC1 biological roles have been limited initially by the laborious purification of the protein from T. gondii tachyzoites and the low yields verified. Then Escherichia coli has been transformed with a construct containing the MIC1 gene, and the obtained recombinant MIC1 (rMIC1) has been purified from the inclusion bodies. Herein, we detail the methodology of heterologous production and purification of rMIC1 and protocols to assay the rMIC1 lectin ability.
