title: Endocytosis of pulchellin and its recombinant B-chain into K-562 cells: binding and uptake studies.
abstract: Most of the type 2 ribosome-inactivating proteins (RIPs) are toxins formed by an RNA-N-glycosidase A-chain polypeptide linked to a lectin B-chain by a single disulfide bond. Members of this protein class vary greatly in cytotoxity, correlating more with B-chain diversity rather than to A-chain differences. Pulchellin is a type 2 ribosome-inactivating protein toxin found in the seeds of Abrus pulchellus tenuiflorus. Recombinant pulchellin B-Chain (rPBC) has been previously produced as inclusion bodies in Escherichia coli and successfully refolded recovering biological activity. New approaches for using this kind of protein as a biotechnological tool require a better understanding of cell targeting, binding, uptake, intracellular routing and delivery. In this work, cell adhesion experiments were used to determine the interaction of rPBC with mammalian cells. Fluorescence and confocal microscopy revealed the intracellular localization and trafficking. Subcellular sorting of the native pulchellin could also be determined. The results support that the endosomal internalization pathway and the retrograde transport through the Golgi apparatus might be used by both native protein and rPBC.
authors.affiliation: Centro de Biotecnologia Molecular Estrutural, Instituto de Física de São Carlos, Universidade de São Paulo, São Carlos, SP, Brazil.
authors.forename: Leandro S
authors.initials: LS
authors.lastname: Goto
authors.forename: Priscila V
authors.initials: PV
authors.lastname: Castilho
authors.forename: Márcia R
authors.initials: MR
authors.lastname: Cominetti
authors.forename: Heloísa S
authors.initials: HS
authors.lastname: Selistre-Araújo
authors.forename: Ana Paula
authors.initials: AP
authors.lastname: Ulian Araújo
mesh_terms: D000818:Animals; D004705:Endocytosis; D006801:Humans; D020014:K562 Cells; D051379:Mice; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism; D054790:Ribosome Inactivating Proteins, Type 2 / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D054790:Ribosome Inactivating Proteins, Type 2