Open AccessBinding and Inhibition of Spermidine Synthase from Plasmodium falciparum and Implications for In Vitro Inhibitor Testing
Author(s) -
Janina Sprenger,
Jannette Carey,
Bo Svensson,
Verena Wengel,
Lo Persson
Publication year - 2016
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0163442
Subject(s) - isothermal titration calorimetry , spermidine , putrescine , plasmodium falciparum , ic50 , biochemistry , binding site , spermine , enzyme , biology , chemistry , stereochemistry , in vitro , malaria , immunology
The aminopropyltransferase spermidine synthase (SpdS) is a promising drug target in cancer and in protozoan diseases including malaria. Plasmodium falciparum SpdS ( Pf SpdS) transfers the aminopropyl group of decarboxylated S-adenosylmethionine (dcAdoMet) to putrescine or to spermidine to form spermidine or spermine, respectively. In an effort to understand why efficient inhibitors of Pf SpdS have been elusive, the present study uses enzyme activity assays and isothermal titration calorimetry with verified or predicted inhibitors of Pf SpdS to analyze the relationship between binding affinity as assessed by K D and inhibitory activity as assessed by IC 50 . The results show that some predicted inhibitors bind to the enzyme with high affinity but are poor inhibitors. Binding studies with Pf SpdS substrates and products strongly support an ordered sequential mechanism in which the aminopropyl donor (dcAdoMet) site must be occupied before the aminopropyl acceptor (putrescine) site can be occupied. Analysis of the results also shows that the ordered sequential mechanism adequately accounts for the complex relationship between IC 50 and K D and may explain the limited success of previous efforts at structure-based inhibitor design for Pf SpdS. Based on Pf SpdS active-site occupancy, we suggest a classification of ligands that can help to predict the K D −IC 50 relations in future design of new inhibitors. The present findings may be relevant for other drug targets that follow an ordered sequential mechanism.