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Cloning, expression and purification of enzyme CRK3 from Leishmania mexicana (LmCRK3) and cycline 6 Leishmania major (LmCYC6)

Grant number: 19/01678-7
Support type:Scholarships in Brazil - Scientific Initiation
Effective date (Start): May 01, 2019
Effective date (End): April 30, 2020
Field of knowledge:Biological Sciences - Biochemistry
Principal Investigator:Wagner Alves de Souza Júdice
Grantee:Fábio da Silva Siqueira
Home Institution: Pró-Reitoria Acadêmica. Universidade de Mogi das Cruzes (UMC). Campus da Sede Mogi das Cruzes. Mogi das Cruzes , SP, Brazil

Abstract

Leishmaniasis are pathologies considered neglected by the World Health Organization, approximately 70 countries are part of the endemic region of the disease, but it is worth mentioning that the most affected are tropical and subtropical. Although developed countries report cases, underdeveloped or developing countries are the most affected due to poor health status, since the females of the vectors use accumulation of organic material and humid places for egg laying. Currently about 2 million people carry the parasite, and an average of 500 thousand new cases are registered each year. Current pharmacological treatments have high toxicity and may take more than a half-year, so new leishmanicidal drugs are under study. It is known that CRK3 is essential for the progression of the parasite's cell cycle, which has made it a major pharmacological target. The present project aims at the cloning, expression and purification of the enzymes LmCRK3, LdCRK3 and the cyclin LmjCYC6. Cloning will be performed on E. coli dH5± strain and expression in E. coli BL21 strain with IPTG induction. Purification will be done by affinity chromatography on a nickel resin column and the confirmation of the target proteins will be by Western blot technique. The obtaining of these enzymes will be important so that in later studies it is possible the discovery of new molecules inhibitors.