Isolation and intracellular localization of insulin-like proteins from leaves of Bauhinia variegata

• Main Authors: C.R. Azevedo, F.M. Maciel, L.B. Silva, A.T.S. Ferreira, M. da Cunha, O.L.T. Machado, K.V.S. Fernandes, A.E.A. Oliveira, J. Xavier-Filho
• Format: Article
• Language: English
• Published: Associação Brasileira de Divulgação Científica 2006-11-01
• Series: Brazilian Journal of Medical and Biological Research
• Subjects: Bauhinia variegata; Plant insulin; Chloroplast; Medicinal plant
• Online Access: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2006001100007

Evidence based on immunological cross-reactivity and anti-diabetic properties has suggested the presence of insulin-like peptides in plants. The objective of the present study was to investigate the presence of insulin-like proteins in the leaves of Bauhinia variegata ("pata-de-vaca", "mororó"), a plant widely utilized in popular medicine as an anti-diabetic agent. We show that an insulin-like protein was present in the leaves of this plant. A chloroplast protein with a molecular mass similar to that of bovine insulin was extracted from 2-mm thick 15% SDS-PAGE gels and fractionated with a 2 x 24 cm Sephadex G-50 column. The activity of this insulin-like protein (0.48 mg/mL) on serum glucose levels of four-week-old Swiss albino (CF1) diabetic mice was similar to that of commercial swine insulin used as control. Further characterization of this molecule by reverse-phase hydrophobic HPLC chromatographic analysis as well as its antidiabetic activity on alloxan-induced mice showed that it has insulin-like properties. Immunolocalization of the insulin-like protein in the leaves of B. variegata was performed by transmission electron microscopy using a polyclonal anti-insulin human antibody. Localization in the leaf blades revealed that the insulin-like protein is present mainly in chloroplasts where it is also found associated with crystals which may be calcium oxalate. The presence of an insulin-like protein in chloroplasts may indicate its involvement in carbohydrate metabolism. This finding has strengthened our previous results and suggests that insulin-signaling pathways have been conserved through evolution.