Characterization of thiamine pyrophosphokinase of vitamin B1 biosynthetic pathway as a drug target of Leishmania donovani

Abstract

Vitamin B1 is an essential cofactor for enzymes involved in the metabolism of carbohydrates, particularly Transketolases. These enzymes are amenable to therapeutic interventions because of their specificity. In the final step of the Vitamin B1 biosynthesis pathway, Thiamine Pyrophosphokinase (TPK) converts thiamin into its active form, Thiamin Pyrophosphate (TPP), allowing researchers to investigate the functional importance of this enzyme and the pathway’s dispensability in Leishmania donovani, a protozoan parasite that causes visceral leishmaniasis. In this study, various in silico, biochemical, biophysical, and cellular assays-based experiments have been conducted to identify and characterize LdTPK, and to provide a sound platform for the discovery of potential LdTPK inhibitors. LdTPK structural modelling ensured high protein quality. Oxythiamine and pyrithiamine were found to bind well with LdTPK with considerable binding energies, and MD simulation-based experiments indicated the stability of the complexation. Additionally, LdTPK1 was found to activate ROS defense in amastigotes, and its inhibition using oxythiamine and pyrithiamine led to the growth inhibition of L. donovani promastigotes and intracellular amastigotes. These findings highlight LdTPK as a promising target for the development of new anti-leishmanial agents. An in-depth analysis of the enzymes involved in TPP biosynthesis in L. donovani has the potential to yield novel therapeutic strategies for Leishmaniasis.

Communicated by Ramaswamy H. Sarma

Keywords:

• Leishmania donovani
• vitamin B1
• thiamine pyrophosphokinase
• thiamine
• inhibition

Acknowledgements

S. Singh is a recipient of the Innovative Young Biotechnologist Award (IYBA) and National Bioscientist Award from Department of Biotechnology (DBT). Authors acknowledge Central Instrument Facility of School of life Sciences, Special Centre for Molecular Medicine and Advance Instrument research Facility, JNU for various instruments. RRK is thankful to Prof. Rentala Madhubala (superannuated), School of Life Sciences, JNU for allowing her research laboratory to conduct few experiments. All authors acknowledge University Grant Commission (UGC) for the support. RJ acknowledges UGC for doctoral research fellowship (award letter sr. no.: 2061430573; ref. no.: 22/06/2014(i)EU-V).

Disclosure statement

No potential conflict of interest was reported by the authors.

Data availability statement

The datasets used and/or analysed in the current study are available from the corresponding author on reasonable request.

Author contributions

RRK, VS and S. Singh contributed to the study conception and design. Material preparation, data collection and analysis were performed by RRK, RJ, VS and S. Singh. AG and RRK performed molecular dynamics simulation studies. The first draft of the manuscript was written by RRK and RJ. SA and NP contributed to the manuscript writing. All authors read and approved the final manuscript.

Additional information

Funding

This work was supported by funding from Science and Engineering Research Board (SERB) (IPA/2020/000007) and Drug and Pharmaceuticals Research Programe (DPRP) (Project No. P/569/2016-1/TDT) to S. Singh.