Biochemical and structural characterization of the RT domain of Leishmania sp. telomerase reverse transcriptase

Author(s)

da Silva, Vitor Luiz ; de Paiva, Stephany Cacete ; de Oliveira, Hamine Cristina ; Fernandes, Carlos Alexandre H. ; Salvador, Guilherme Henrique Marchi ; Fontes, Marcos Roberto de M. ; Cano, Maria Isabel N.

• Document type: Journal article
• Source: BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS; v. 1867, n. 11, p. 10-pg., 2023-09-28.

Abstract

Background: The Leishmania genus comprises parasites that cause leishmaniasis, a neglected disease spread worldwide. Leishmania sp. telomeres are composed of TTAGGG repeats maintained by telomerase. In most eukaryotes, the enzyme minimal complex contains the TER (telomerase RNA) and the TERT (telomerase reverse transcriptase) components. The TERT holds the enzyme catalytic core and is formed by four structural and functional domains (TEN, Telomerase Essential N-terminal; TRBD, Telomerase RNA Binding Domain; RT, the reverse transcriptase domain and CTE, C-Terminal Extension domain). Methods and results: Amino acid sequence alignments, protein structure prediction analysis, and protein: nucleic acid interaction assays were used to show that the Leishmania major RT domain preserves the canonical structural elements found in higher eukaryotes, including the canonical motifs and the aspartic acid residues that stabilize the Mg2+ ion cofactor. Furthermore, amino acid substitutions specific to the Leishmania genus and partial conservation of the residues involved with nucleic acid interactions are shown. The purified recombinant Leishmania RT protein is biochemically active and interacts with the G-rich telomeric strand and the TER template sequence. Conclusion: Our results highlight that the telomerase catalysis mechanism is conserved in a pathogen of medical importance despite the structural peculiarities present in the parasite's RT domain. (AU)

• FAPESP's process: 19/05958-4 - Multi-User Equipment approved in grant 2013/24705-3: high pressure liquid chromatography system
• Grantee: Marcos Roberto de Mattos Fontes
• Support Opportunities: Multi-user Equipment Program
• FAPESP's process: 21/14798-0 - Functional and structural studies of the PinX1 orthologue and its influence on Leishmania spp telomeres
• Grantee: Stephany Cacete de Paiva
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)
• FAPESP's process: 18/04375-2 - Studies about the biogenesis and composition of the Leishmania spp. ribonucleoprotein complex and its regulation
• Grantee: Maria Isabel Nogueira Cano
• Support Opportunities: Research Projects - Thematic Grants
• FAPESP's process: 19/05239-8 - Comparative study between different biophysical techniques for further understanding of the nuclear importation mechanism
• Grantee: Hamine Cristina de Oliveira
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)