PATHOGENESIS MICROBIAL Microbial Pathogenesis 44 (2008) 329–338 Trypanosoma cruzi heparin-binding proteins and the nature of the host cell heparan sulfate-binding domain Francisco Odencio Rodrigues de Oliveira Jr. a , Carlos Roberto Alves b , Cla´udia Magalha˜es Calvet a , Leny Toma c , Rodrigo Ippolito Bouc - as c , Helena Bociani Nader c , Luzia Monteiro de Castro Coˆrtes b , Marco Aure´lio Krieger d , Maria de Nazareth S.L. Meirelles a , Mirian Claudia de Souza Pereira a,Ã a Laborato´rio de Ultra-estrutura Celular, IOC/FIOCRUZ, Av. Brasil 4365, Manguinhos, 21040-900 Rio de Janeiro, RJ, Brazil b Laborato ´rio de Biologia Molecular e Doenc - as Endeˆmicas, IOC/FIOCRUZ, RJ, Brazil c Departamento de Bioquı´mica, Universidade Federal de Sa˜o Paulo, UNIFESP, SP, Brazil d Instituto de Biologia Molecular do Parana´, IBMP, FIOCRUZ, PR, Brazil Received 31 July 2007; received in revised form 15 October 2007; accepted 16 October 2007 Available online 22 October 2007 Abstract Trypanosoma cruzi invasion is mediated by receptor–ligand recognition between the surfaces of both parasite and target cell. We have previously demonstrated the role of heparan sulfate proteoglycan in the attachment and invasion of T. cruzi in cardiomyocytes. Herein, we have isolated the T. cruzi heparin-binding proteins (HBP-Tc) and investigated the nature of cardiomyocyte heparan sulfate (HS)- binding site to the parasite surface ligand. Two major heparin-binding proteins with molecular masses of 65.8 and 59 kDa were observed in total extract of amastigote and trypomastigote forms of T. cruzi. Hydrophobic [S 35 ]methionine labeled proteins eluted from heparin–sepharose affinity chromatography also revealed both proteins in trypomastigotes but only the 59 kDa is strongly recognized by biotin-conjugated glycosaminoglycans. Competition assays were performed to analyze the role of sulfated proteoglycans, including heparin, keratan sulfate and both acetylated and highly sulfated domains of heparan sulfate, in the recognition and invasion process of T. cruzi. Significant inhibitions of 84% and 35% in the percentage of infection were revealed after treatment of the parasites with heparin and the N-acetylated/ N-sulfated heparan sulfate domain, respectively, suggesting the important role of the glycuronic acid and NS glucosamine domain of the HS chain in the recognition of the HBP-Tc during the T. cruzi–cardiomyocyte interaction. r 2007 Elsevier Ltd. All rights reserved. Keywords: Trypanosoma cruzi; Cardiomyocytes; Glycosaminoglycans; Recognition process; Chromatography 1. Introduction Chagas’ disease, caused by the parasite Trypanosoma cruzi, is an important cause of irreversible cardiomyopathy which affects approximately 13 million people in Latin America [1]. In addition, this infection poses a potential hazard to many countries including those in North America and Europe because of blood transfusion and organ transplantation [2,3]. The pathogenesis of chronic Chagas’ disease is not completely understood and may be related to parasite persistence, parasite antigens [4,5] and/or autoimmune mechanisms [6–8]. Invasion of mam- malian cells by T. cruzi is mediated by receptor–ligand recognition between the surfaces of both parasite and target cells. Carbohydrate-binding protein in the parasite [9,10] and host cell galactose, manose, N-acetylgalactosa- mine residues are important for attachment and host cell invasion [11–13]. The parasite expresses a family of active and inactive trans-sialidase, a glycosylphosphatidy- linositol anchored protein on its surface which has been demonstrated to be involved in host cell invasion and immunomodulation [14,15]. In addition to parasite ligands, several host cell surface molecules have been implicated ARTICLE IN PRESS www.elsevier.com/locate/micpath 0882-4010/$ - see front matter r 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.micpath.2007.10.003 Ã Corresponding author. Tel.: +55 21 2598 4331; fax: +55 21 2260 4434. E-mail address: mirian@ioc.fiocruz.br (M.C. Souza Pereira).