The single cyclic nucleotide-specific phosphodiesterase of the intestinal parasite Giardia lamblia represents a potential drug target.

Stefan Kunz,Geert Jan Sterk,Vreni Balmer,Michael P Pollastri,Norbert Müller,Cornelia Spycher,Andrew Hemphill,Rob Leurs,Aaron R Jex

doi:10.1371/journal.pntd.0005891

Stefan Kunz, Geert Jan Sterk + Show 7 more

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https://doi.org/10.1371/journal.pntd.0005891

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Abstract

BackgroundGiardiasis is an intestinal infection correlated with poverty and poor drinking water quality, and treatment options are limited. According to the Center for Disease Control and Prevention, Giardia infections afflict nearly 33% of people in developing countries, and 2% of the adult population in the developed world. This study describes the single cyclic nucleotide-specific phosphodiesterase (PDE) of G. lamblia and assesses PDE inhibitors as a new generation of anti-giardial drugs.MethodsAn extensive search of the Giardia genome database identified a single gene coding for a class I PDE, GlPDE. The predicted protein sequence was analyzed in-silico to characterize its domain structure and catalytic domain. Enzymatic activity of GlPDE was established by complementation of a PDE-deficient Saccharomyces cerevisiae strain, and enzyme kinetics were characterized in soluble yeast lysates. The potency of known PDE inhibitors was tested against the activity of recombinant GlPDE expressed in yeast and against proliferating Giardia trophozoites. Finally, the localization of epitope-tagged and ectopically expressed GlPDE in Giardia cells was investigated.ResultsGiardia encodes a class I PDE. Catalytically important residues are fully conserved between GlPDE and human PDEs, but sequence differences between their catalytic domains suggest that designing Giardia-specific inhibitors is feasible. Recombinant GlPDE hydrolyzes cAMP with a Km of 408 μM, and cGMP is not accepted as a substrate. A number of drugs exhibit a high degree of correlation between their potency against the recombinant enzyme and their inhibition of trophozoite proliferation in culture. Epitope-tagged GlPDE localizes as dots in a pattern reminiscent of mitosomes and to the perinuclear region in Giardia.ConclusionsOur data strongly suggest that inhibition of G. lamblia PDE activity leads to a profound inhibition of parasite proliferation and that GlPDE is a promising target for developing novel anti-giardial drugs.

Highlights

Giardia lamblia is a protozoan parasite that causes giardiasis, an intestinal disease with symptoms such as diarrhea, nausea, and malabsorption [1]
Important residues are fully conserved between gene coding for a class I PDE (GlPDE) and human PDEs, but sequence differences between their catalytic domains suggest that designing Giardia-specific inhibitors is feasible
Our data strongly suggest that inhibition of G. lamblia PDE activity leads to a profound inhibition of parasite proliferation and that GlPDE is a promising target for developing novel antigiardial drugs

Summary

Introduction

Giardia lamblia is a protozoan parasite that causes giardiasis, an intestinal disease with symptoms such as diarrhea, nausea, and malabsorption [1]. Trophozoites are the disease-causing stage and colonize the upper small intestine of humans and other vertebrates. They form cysts, which are shed into the environment via the fecal route and which are orally transmitted, mostly via contaminated water. Giardiasis occurs worldwide, predominantly in resource-poor countries with low standards of sanitation, and represents a major cause of non-bacterial diarrhea with 280 million symptomatic human cases every year [2]. Giardiasis is an intestinal infection correlated with poverty and poor drinking water quality, and treatment options are limited. This study describes the single cyclic nucleotidespecific phosphodiesterase (PDE) of G. lamblia and assesses PDE inhibitors as a new generation of anti-giardial drugs

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