Biochemical characterization of the bifunctional enzyme dihydrofolate reductase-thymidylate synthase from Leishmania (Viannia) and its evaluation as a drug target
Abstract
Introduction: Dihydrofolate reductase (DHFR) has been used successfully as a drug target in the area of anti-bacterial, anti-cancer and anti-malarial therapy. Although this bifunctional enzyme is also a potential drug target for treatment of leishmaniasis, there have been no reports on its efficacy against Leishmania (Viannia) species.
Materials and methods: The gene encoding the bifunctional DHFR and thymidylate synthase (TS) of Le. (V.) braziliensis was isolated and expressed in E. coli. The enzyme was purified and characterized. The inhibitory effects of antifolates and four aporphine alkaloids on its activity were evaluated.
Results: The full-length gene consists of a 1560-bp open reading frame encoding a 58 kDa translated peptide containing DHFR and TS domains linked together in a single polypeptide chain. The recombinant DHFR-TS enzyme revealed Km and Vmax values of 55.35 ± 4.02 μM (mean ± SE) and 0.02 ± 5.34 x10-4 μM/min respectively for dihydrofolic acid (H2F). The Le. braziliensis rDHFR-TS have Ki values for antimicrobial antifolates in the µM range. Methotrexate (MTX) was a more-potent inhibitor of enzymatic activity (Ki = 22.0 µM) than trimethoprim (Ki = 33 µM) and pyrimethamine (Ki = 68 µM). These Ki values are significantly lower than those obtained for the aporphine alkaloids.
Conclusion: The results of the study show the inhibitory effect of antifolate drugs on enzymatic activity,indicating that Le. braziliensis rDHFR-TS could be a model to studying antifolate compounds aspotential antiprotozoal drugs.
doi: http://dx.doi.org/10.7705/biomedica.v33i3.1434
Downloads
Some similar items:
- Erika Santamaría, Mónica Castillo, Rocío Cárdenas, Felio Bello, Martha Ayala, Cristina Ferro, Vectorial cornpetence of Lutzomyia species from the verrucarum (Diptera, Psychodidae) group found in a Leishmania braziliensis endernic focus in Reventones, Cundinamarca , Biomedica: Vol. 19 No. 2 (1999)
- Camilo Andrés Morales, Juliana Palacio, Gerzaín Rodríguez, Yenny Carolina Camargo, Zosteriform cutaneous leishmaniasis due to Leishmania (Viannia) panamensis and Leishmania (Viannia) braziliensis: Report of three cases , Biomedica: Vol. 34 No. 3 (2014)
- Juan Sergio Mollinedo, Zoraida Mollinedo , Marcelo Magne , Wilson J. Gironda, Óscar D. Salomón , Leishmaniasis en Bolivia. Comprehensive review and current status in Tarija, in the border with Argentina , Biomedica: Vol. 40 No. Supl. 1 (2020): Mayo, Infecciones en el trópico
- Caroline de Souza Silverio , Carolina Bonilla, Vitamin D-associated genetic variants in the Brazilian population: Investigating potential instruments for Mendelian randomization , Biomedica: Vol. 44 No. 1 (2024)
Article metrics | |
---|---|
Abstract views | |
Galley vies | |
PDF Views | |
HTML views | |
Other views |