Cloning, expression and characterization of HtrA proteins of Leptospira interrogans

• Grant number: 17/01411-5
• Support Opportunities: Scholarships in Brazil - Master
• Start date: May 01, 2017
• End date: February 28, 2019
• Field of knowledge: Biological Sciences - Microbiology
• Agreement: Coordination of Improvement of Higher Education Personnel (CAPES)
• Principal Investigator: Ana Lucia Tabet Oller do Nascimento
• Grantee: Brenda Daroz
• Host Institution: Instituto Butantan. Secretaria da Saúde (São Paulo - Estado). São Paulo , SP, Brazil
• Associated research grant: 14/50981-0 - Search for surface proteins among the genome sequences of Leptospira interrogans: functional and immunological characterization to understanding mechanisms involved in the bacterial pathogenesis, AP.TEM

Abstract

Leptospirosis is a zoonosis of global importance recently included in the list of Neglected Tropical Diseases, caused by pathogenic bacteria of the genus Leptospira. Rodents play the major reservoirs of the disease by allowing the colonization of leptospires in the proximal renal tubules and excreting them alive in the urine. According to the World Health Organization, the annual number of clinical cases in the world remains underestimated and there is no effective vaccine so far. The identification of conserved outer membrane proteins among pathogenic strains are the main research targets to elucidate pathogenicity mechanisms. It is these proteins that are involved in the interaction of the bacteria with the external environment, and can also serve as a target for the immune system. Some spirochetes utilize outer membrane proteases as a mechanism for dissemination in the tissue of the host. Thus, the project proposes the cloning and expression of two proteins predicted to be type HtrA identified by bioinformatics in the genome of Leptospira interrogans. The genes encoding these proteins, LIC11111 and LIC1112, will be amplified from the genomic DNA and the cloned insert into expression vector in E. coli. Recombinant proteins will be expressed with a sequence of six histidine residues in the N-terminal region for purification by metal affinity chromatography. It will be evaluated: (a) the immunogenicity activity of proteins in animal model, (b) reactivity of the recombinant proteins to serum from patients diagnosed with leptospirosis, and (c) interaction of these proteins with host macromolecules and their proteolytic potential on them. (AU)

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

NASCIMENTO FILHO, EDSON G.; VIEIRA, MONICA L.; TEIXEIR, ALINE F.; SANTOS, JADEMILSON C.; FERNANDES, LUIS G. V.; PASSALIA, FELIPE J.; DAROZ, BRENDA B.; ROSSINI, AMANDA; KOCHI, LEANDRO T.; CAVENAGUE, MARIA F.; et al. Proteomics as a tool to understand Leptospira physiology and virulence: Recent advances, challenges and clinical implications. JOURNAL OF PROTEOMICS, v. 180, n. SI, p. 80-87, MAY 30 2018. (17/01411-5, 17/06731-8, 14/50981-0, 17/01102-2, 16/11541-0, 16/04295-3, 16/01384-5)

TEIXEIRA, ALINE F.; FERNANDES, V, LUIS G.; CAVENAGUE, MARIA F.; TAKAHASHI, MARIA B.; SANTOS, JADEMILSON C.; PASSALIA, FELIPE J.; DAROZ, BRENDA B.; KOCHI, LEANDRO T.; VIEIRA, MONICA L.; NASCIMENTO, ANA L. T. O.. Adjuvanted leptospiral vaccines: Challenges and future development of new leptospirosis vaccines. Vaccine, v. 37, n. 30, p. 3961-3973, JUL 9 2019. (17/01411-5, 17/06731-8, 17/25167-6, 14/50981-0, 17/01102-2, 16/11541-0, 17/26223-7, 17/00236-5, 16/01384-5)

NASCIMENTO FILHO, EDSON G.; VIEIRA, MONICA L.; TEIXEIR, ALINE F.; SANTOS, JADEMILSON C.; FERNANDES, LUIS G. V.; PASSALIA, FELIPE J.; DAROZ, BRENDA B.; ROSSINI, AMANDA; KOCHI, LEANDRO T.; CAVENAGUE, MARIA F.; et al. Proteomics as a tool to understand Leptospira physiology and virulence: Recent advances, challenges and clinical implications. JOURNAL OF PROTEOMICS, v. 180, p. 8-pg., 2018-05-30. (14/50981-0, 17/01411-5, 17/01102-2, 16/01384-5, 16/04295-3, 16/11541-0, 17/06731-8)