Leishmania tarentolae is a non-pathogenic trypanosomatid isolated from lizards widely used for heterologous protein expression and extensively studied to understand the pathogenic mechanisms of leishmaniasis. The repertoire of leishmanolysin genes was reported to be expanded in L. tarentolae genome, but no proteolytic activity was detected. Here, we analyzed L. tarentolae leishmanolysin proteins from the genome to the structural levels and evaluated the enzymatic activity of the wild-type and overexpressing mutants of leishmanolysin. A total of 61 leishmanolysin sequences were retrieved from the L. tarentolae genome. Five of them were selected for phylogenetic analysis, and for three of them, we built 3D models based on the crystallographic structure of L. major ortholog. Molecular dynamics simulations of these models disclosed a less negative electrostatic potential compared to the template. Subsequently, L. major LmjF.10.0460 and L. tarentolae LtaP10.0650 leishmanolysins were cloned in a pLEXSY expression system into L. tarentolae. Proteins from the wild-type and the overexpressing parasites were submitted to enzymatic analysis. Our results revealed that L. tarentolae leishmanolysins harbor a weak enzymatic activity about three times less abundant than L. major leishmanolysin. Our findings strongly suggest that the less negative electrostatic potential of L. tarentolae leishmanolysin can be the reason for the reduced proteolytic activity detected in this parasite.

Coleção de Protozários da Fundação Oswaldo Cruz Rio de Janeiro 21040 900 Brazil

Grupo de Biofísica Computacional e Modelagem Molecular Programa de Computação Científica Rio de Janeiro 21040 360 Brazil

Laboratório de Biologia Computacional e Sistemas Instituto Oswaldo Cruz Fundação Oswaldo Cruz Rio de Janeiro 21040 360 Brazil

Laboratório de Estudos Avançados de Microrganismos Emergentes e Resistentes Instituto de Microbiologia Paulo de Góes Universidade Federal do Rio de Janeiro Rio de Janeiro 21941 902 Brazil

Laboratório de Estudos Integrados em Protozoologia Instituto Oswaldo Cruz Fundação Oswaldo Cruz Rio de Janeiro 21040 360 Brazil

Laboratório de Genômica Funcional e Bioinformática Instituto Oswaldo Cruz Fundação Oswaldo Cruz Rio de Janeiro 21040 360 Brazil

Life Science Research Centre Faculty of Science University of Ostrava 70200 Ostrava Czech Republic

Martsinovsky Institute of Medical Parasitology Tropical and Vector Borne Diseases Sechenov University 119435 Moscow Russia

Programa de Pós Graduação em Bioquímica Instituto de Química Universidade Federal do Rio de Janeiro Rio de Janeiro 21941 902 Brazil

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