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Master's Dissertation
DOI
https://doi.org/10.11606/D.87.2016.tde-24082016-094832
Document
Author
Full name
Lucas Pereira e Silva
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2016
Supervisor
Committee
Nascimento, Ana Lucia Tabet Oller do (President)
Astray, Renato Mancini
Lima, Maria Regina D'Imperio
Title in Portuguese
Clonagem, expressão e caracterização de prováveis proteínas de membrana indentificadas no genoma de Leptospira interrogans.
Keywords in Portuguese
Leptospira
Leptospirose
Proteínas recombinantes
Abstract in Portuguese
A leptospirose é uma doença sistêmica, causada por bactérias patogênicas do gênero Leptospira. O desenvolvimento de novas estratégias para prevenir a doença é necessário. As pesquisas atuais têm interesse em identificar antígenos conservados que estão envolvidos nas interações patógeno-hospedeiro. Dois genes de L. interrogans foram selecionados, clonados, expressos e suas respectivas proteínas caracterizadas. Os genes foram amplificados por PCR e clonados no vetor de expressão PAE. As proteínas recombinantes foram purificadas por cromatografia de afinidade ao metal. A proteína rLIC10821 foi capaz de se ligar a laminina, plasminogênio e fibrinogênio. Ambas as proteínas foram localizadas na membrana externa de acordo com as três metodologias utilizadas: imunofluorescência, proteinase K, leptospira intacta. A proteína rLIC10821 que interagiu com o PLG foi capaz de gerar plasmina. Após a interação da proteína rLIC10821 com o fibrinogênio, foi possível identificar uma diminuição de 60% no coágulo de fibrina.
Title in English
Cloning, expression and characterization of probable membrane proteins identified on the Leptospira interrogans genome.
Keywords in English
Leptospira
Leptospirosis
Recombinant proteins
Abstract in English
Leptospirosis is a systemic disease caused by pathogenic bacteria of genus Leptospira. The development of new strategies to prevent the disease is needed. Currently research has focused to identify conserved antigens related to the host-pathogen interaction. Two genes of L. interrogans were selected, cloned, expressed and its proteins characterized. The genes were amplified by PCR and cloned into expression vetor pAE. The recombinant proteins were purified by chromatography of metal affinity. The protein rLIC10821 were able to bind to laminin, plasminogen and fibrinogen. Both proteins were localized in the outer membrane according three methodologies: immunofluorescence, proteinase K, intact leptospira. The protein rLIC10821 interacted with PLG was able to generate plasmin. After the interaction of the protein rLIC10821 with fibrinogen, we could identify a decrease of 60% in the fibrin clot.
 
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Publishing Date
2016-08-24
 
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