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

• Published in: Biochimica et biophysica acta. General subjects Vol. 1867; no. 11; p. 130451
• Main Authors: 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.
• Format: Journal Article
• Language: English
• Published: Elsevier B.V 01.11.2023
• Subjects: amino acid sequences; aspartic acid; catalytic activity; eukaryotic cells; Leishmania; Leishmania major; leishmaniasis; parasites; pathogens; prediction; Protein: Nucleic acid interactions; Recombinant protein purification; RNA; RT domain; Structure prediction; telomerase; telomeres; TER; CD; RT; LmTERT; IFD-TRAP; Recombinant protein purification; TRBD; Structure prediction; RT domain; Protein: Nucleic acid interactions; TERT; CTE; Leishmania; LeishTER; TEN
• ISSN: 0304-4165; 1872-8006; 1872-8006
• DOI: 10.1016/j.bbagen.2023.130451

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). 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. 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. •The Leishmania TERT RT domain contains amino acid substitutions specific to the genus.•The main structural components of the RT domain are conserved in Leishmania TERT.•The in silico model showed that the Leishmania TERT RT preserves the canonical motifs.•The Leishmania RT domain presents the Asp residues that stabilize the Mg2+ ion cofactor.•The Leishmania RT domain is active and interacts with the telomeric DNA and LeishTER.