Magnesium-Dependent Ecto-ATP Diphosphohydrolase Activity in Herpetomonas muscarum muscarum

Abstract

In the present work we characterized the ecto-ATP diphosphohydrolase activity of the trypanosomatid parasite Herpetomonas muscarum muscarum. This parasite hydrolyzed ATP at a rate of 15.52 nmol Pi/mg protein/min and this activity reached a maximum at pH 7.5. Classical inhibitors of acid phosphatases, such as sodium orthovanadate (N_aVO₃), sodium fluoride (NaF), and ammonium molybdate presented no effect on this activity. MgCl₂, ZnCl₂, and MnCl₂ stimulated the ATP hydrolysis by H. m. muscarum. The ecto-ATPase activity was insensitive to oligomycin and sodium azide, two inhibitors of mitochondrial Mg-ATPase, bafilomycin A₁, a V-ATPase inhibitor, ouabain, a Na⁺+K⁺-ATPase inhibitor and to levamizole, an inhibitor of alkaline phosphatase. An extracellular impermeant inhibitor 4,4′-diisothiocyanostylbene 2′,2′-disulfonic acid (DIDS) and a inhibitor of some ecto-ATPases, suramin, which is also a competitive antagonist of P₂-purinergic receptors, promoted a great inhibition on the ATP hydrolysis. This enzyme is able to hydrolysis ATP, ADP, UTP, and UDP, but not GTP, GDP, CTP, or CDP. ADP inhibited the enzymatic activity in a concentration dependent manner, reaching 70% inhibition.