Characterization of plasma membrane bound inorganic pyrophosphatase from Leishmania donovani promastigotes and amastigotes

S.S Sen, N.R Bhuyan, T Bera

Abstract

Background: Currently, a major problem in the management of visceral leishmaniasis or kala-azar, especially in the Indian subcontinent, is the growing unresponsiveness to conventional antimonial therapy. Membrane bound pyrophophatase (PPases) do not exist in plasma membrane from mammals. Thus, H+-PPases from Leishmania plasma membrane might be potential target in rational chemotherapy of the disease caused by Leishmania parasites.

Objective: To characterize the activities of inorganic pyrophophatase (PPase) in the plasma membrane of Leishmania donovani promastigote and amastigote.

Methods: Culture method of promastigote and amastigote were developed. We assayed PPase activity in isolated plasma membrane of L. donovani.

Results: We characterized K+-PPase present in the plasma membrane of Leishmania donovani and investigated its possible role in the survival of promastigote and amastigote. PPase activity was stimulated by K+ ions and sodium orthovanadate, inhibited by pyrophosphate analogs imidodiphosphate and alendronate, KF, DCCD, thiol reagent parachloromercuribenzenesulfonate (PCMBS), N-ethylmaliemide (NEM), phenylarsineoxide, ABC superfamily transport modulator verapamil and was also by F1Fo-ATPase inhibitor quercetin.

Conclusion: We conclude that there are significant differences within promastigote, amastigote and mammalian host in cytosolic pH homeostasis to merit the inclusion of PPase transporter as putative targets for rational drug design.