title: Intermediate and C-terminal regions of leptospiral adhesin Lsa66 are responsible for binding with plasminogen and extracellular matrix components.
abstract: Leptospirosis, a worldwide zoonotic infection, is an important human and veterinary health problem. We have previously identified a leptospiral multipurpose adhesin, Lsa66, capable of binding extracellular matrix (ECM) components and plasminogen (PLG). In this work, we report the cloning, expression, purification and characterization of three fragments derived from the full-length Lsa66: N-terminal, intermediate and C-terminal regions. We employed Escherichia coli BL21-SI as expression cells. The recombinant fragments tagged with N-terminal His6 were purified by metal-charged chromatography to major protein bands that were recognized by anti-His-tag mAbs. The recombinant fragments were evaluated for their capacity to attach to ECM components and to PLG. The intermediate region bound to laminin, plasma fibronectin and PLG. Laminin also bound to the C-terminal region. Antibodies in leptospirosis-positive serum samples recognized Lsa66, being the immune epitopes located at the N-terminal and intermediate fragments. The data confirm that Lsa66 is expressed during infection and that this protein might have a role in bacterial infection.
authors.affiliation: Interunidades em Biotecnologia, Instituto de Ciências Biomédicas, USP, Avenida Prof. Lineu Prestes, 1730, 05508-900 São Paulo, SP, Brazil.|Centro de Biotecnologia, Instituto Butantan, Avenida Vital Brazil, 1500, 05503-900 São Paulo, SP, Brazil.
authors.forename: Rosane
authors.initials: R
authors.lastname: Oliveira
authors.affiliation: Interunidades em Biotecnologia, Instituto de Ciências Biomédicas, USP, Avenida Prof. Lineu Prestes, 1730, 05508-900 São Paulo, SP, Brazil.|Centro de Biotecnologia, Instituto Butantan, Avenida Vital Brazil, 1500, 05503-900 São Paulo, SP, Brazil.
authors.forename: Renan F
authors.initials: RF
authors.lastname: Domingos
authors.affiliation: Laboratório de Zoonoses Bacterianas do VPS, Faculdade de Medicina Veterinária e Zootecnia, USP, Avenida Prof. Dr Orlando Marques de Paiva, 87, 05508-270 São Paulo, SP, Brazil.
authors.forename: Zenaide M
authors.initials: ZM
authors.lastname: de Morais
authors.affiliation: Laboratório de Zoonoses Bacterianas do VPS, Faculdade de Medicina Veterinária e Zootecnia, USP, Avenida Prof. Dr Orlando Marques de Paiva, 87, 05508-270 São Paulo, SP, Brazil.
authors.forename: Silvio A
authors.initials: SA
authors.lastname: Vasconcellos
authors.affiliation: Instituto Adolfo Lutz, Laboratório Regional de Santos, Núcleo de Ciências Biomédicas, Rua Silva Jardim, 90, 11015-020, Santos, SP, Brazil.
authors.forename: Ivy J
authors.initials: IJ
authors.lastname: Alves
authors.affiliation: Centro de Bacteriologia, Instituto Adolfo Lutz, Av. Dr Arnaldo, 355, 01246-902 São Paulo, SP, Brazil.
authors.forename: Eliete C
authors.initials: EC
authors.lastname: Romero
authors.affiliation: Interunidades em Biotecnologia, Instituto de Ciências Biomédicas, USP, Avenida Prof. Lineu Prestes, 1730, 05508-900 São Paulo, SP, Brazil.|Centro de Biotecnologia, Instituto Butantan, Avenida Vital Brazil, 1500, 05503-900 São Paulo, SP, Brazil.
authors.forename: Ana L T O
authors.initials: ALTO
authors.lastname: Nascimento
mesh_terms: D018829:Adhesins, Bacterial / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D016326:Extracellular Matrix Proteins / Q000378:metabolism*; D005260:Female; D015870:Gene Expression; D008807:Mice, Inbred BALB C; D010958:Plasminogen / Q000378:metabolism*; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D025941:Protein Interaction Mapping; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D018829:Adhesins, Bacterial; D000907:Antibodies, Bacterial; D016326:Extracellular Matrix Proteins; D011994:Recombinant Proteins; D010958:Plasminogen