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Heterologous expression and biochemical characterization of the recombinant nucleoside triphosphate diphosphohydrolase 2 (LbNTPDase2) from Leishmania (Viannia) braziliensis

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Abstract

Leishmania braziliensis is a pathogenic protozoan parasite that causes American Tegumentary Leishmaniasis (ATL), an important tropical neglected disease. ENTPDases are nucleotidases that hydrolyze intracellular and/or extracellular nucleotides. ENTPDases are known as regulators of purinergic signalling induced by extracellular nucleotides. Leishmania species have two isoforms of ENTPDase, and, particularly, ENTPDase2 seems to be involved in infectivity and virulence. In this study, we conducted the heterologous expression and biochemical characterization of the recombinant ENTPDase2 of L. braziliensis (rLbNTPDase2). Our results show that this enzyme is a canonical ENTPDase with apyrase activity, capable of hydrolysing triphosphate and diphosphate nucleotides, and it is dependent on divalent cations (calcium or magnesium). Substrate specificity was characterized as UDP>GDP>ADP>GTP>ATP=UTP. The enzyme showed optimal activity at a neutral to basic pH and was partially inhibited by suramin and DIDS. Furthermore, the low apparent Km for ADP suggests that the enzyme may play a role in adenosine-mediated signalling. The biochemical characterization of this enzyme can open new avenues for using LbNTPDase2 as a drug target.

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Acknowledgements

The authors gratefully acknowledge the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) for the fellowship granted to JLRF and Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG) for student’s scholarships.

Funding

This study was sponsored in part by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior—Brasil (CAPES)—grant code 001.

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Authors and Affiliations

  1. Biochemistry and Molecular Biology Department, Universidade Federal de Viçosa, Viçosa, MG, Brazil

    Nancy da Rocha Torres Pavione, João Victor Badaró de Moraes, Isadora Cunha Ribeiro, Raissa Barbosa de Castro, Walmir da Silva, Anna Cláudia Alves de Souza, Victor Hugo Ferraz da Silva, Raphael de Souza Vasconcellos, Gustavo da Costa Bressan & Juliana Lopes Rangel Fietto

  2. General Biology Department, Universidade Federal de Viçosa, Viçosa, MG, Brazil

    Nancy da Rocha Torres Pavione & João Victor Badaró de Moraes

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  1. Nancy da Rocha Torres Pavione
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Contributions

All authors contributed to the conception and design of the study. Material preparation, data collection, and analysis were performed by NdRTP. JVBdM, ICR, RBdC, WdS, ACAdS, and VHFdS. The first draft of the manuscript was written by NdRTP; the first revision was done by JLRF, and all authors commented on previous versions of the manuscript. The data and analyses were carefully reviewed by JLRF. All authors read and approved the final manuscript.

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Correspondence to Juliana Lopes Rangel Fietto.

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Nancy da Rocha Torres Pavione declares that she has no conflict of interest.

João Victor Badaró de Moraes declares that he has no conflict of interest.

Isadora Cunha Ribeiro declares that she has no conflict of interest.

Raissa Barbosa de Castro declares that she has no conflict of interest.

Walmir da Silva declares that he has no conflict of interest.

Anna Cláudia Alves de Souza declares that she has no conflict of interest.

Victor Hugo Ferraz da Silva declares that he has no conflict of interest.

Raphael de Souza Vasconcellos declares that he has no conflict of interest.

Gustavo da Costa Bressan declares that he has no conflict of interest.

Juliana Lopes Rangel Fietto declares that she has no conflict of interest.

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da Rocha Torres Pavione, N., de Moraes, J.V.B., Ribeiro, I.C. et al. Heterologous expression and biochemical characterization of the recombinant nucleoside triphosphate diphosphohydrolase 2 (LbNTPDase2) from Leishmania (Viannia) braziliensis. Purinergic Signalling (2023). https://doi.org/10.1007/s11302-023-09980-9

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