• Home
  • Search Results
  • Leishmania amazonensis: characterization of an ecto-pyrophosphatase activity.

Leishmania amazonensis: characterization of an ecto-pyrophosphatase activity.

Experimental parasitology (2013-12-10)
Anita Leocadio Freitas-Mesquita, André Luiz Fonseca-de-Souza, José Roberto Meyer-Fernandes
ABSTRACT

Several ecto-enzymatic activities have been described in the plasma membrane of the protozoan Leishmania amazonensis, which is the major etiological agent of diffuse cutaneous leishmaniasis in South America. These enzymes, including ecto-phosphatases, contribute to the survival of the parasite by participating in phosphate metabolism. This work identifies and characterizes the extracellular hydrolysis of inorganic pyrophosphate related to an ecto-pyrophosphatase activity of the promastigote form of L. amazonensis. This ecto-pyrophosphatase activity is insensitive to MnCl2 but is strongly stimulated by MgCl2. This stimulation was not observed during the hydrolysis of p-nitrophenyl phosphate (p-NPP) or β-glycerophosphate, two substrates for different ecto-phosphatases present in the L. amazonensis plasma membrane. Furthermore, extracellular PPi hydrolysis is more efficient at alkaline pHs, while p-NPP hydrolysis occurs mainly at acidic pHs. These results led us to conclude that extracellular PPi is hydrolyzed not by non-specific ecto-phosphatases but rather by a genuine ecto-pyrophosphatase. In the presence of 5mM MgCl2, the ecto-pyrophosphatase activity from L. amazonensis is sensitive to micromolar concentrations of NaF and millimolar concentrations of CaCl2. Moreover, this activity is significantly higher during the first days of L. amazonensis culture, which suggests a possible role for this enzyme in parasite growth.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Magnesium chloride, anhydrous, ≥98%
Sigma-Aldrich
Magnesium chloride solution, for molecular biology, 1.00 M±0.01 M
Sigma-Aldrich
Calcium chloride dihydrate, ACS reagent, ≥99%
Sigma-Aldrich
Calcium chloride solution, BioUltra, for molecular biology, ~1 M in H2O
Sigma-Aldrich
Sodium fluoride, ACS reagent, ≥99%
Sigma-Aldrich
L-(+)-Tartaric acid, ACS reagent, ≥99.5%
Sigma-Aldrich
Calcium chloride dihydrate, for molecular biology, ≥99.0%
Sigma-Aldrich
Calcium chloride, meets USP testing specifications
Sigma-Aldrich
Calcium chloride dihydrate, BioReagent, suitable for cell culture, suitable for insect cell culture, suitable for plant cell culture, ≥99.0%
Sigma-Aldrich
Manganese(II) chloride tetrahydrate, ReagentPlus®, ≥99%
Sigma-Aldrich
Calcium chloride, anhydrous, powder, 99.99% trace metals basis
Sigma-Aldrich
Calcium chloride, anhydrous, BioReagent, suitable for insect cell culture, suitable for plant cell culture, ≥96.0%
Sigma-Aldrich
Calcium chloride dihydrate, ReagentPlus®, ≥99.0%
Sigma-Aldrich
Manganese(II) chloride tetrahydrate, ACS reagent, ≥98%
Sigma-Aldrich
Magnesium chloride, powder, <200 μm
Sigma-Aldrich
Magnesium chloride solution, BioUltra, for molecular biology, ~1 M in H2O
Sigma-Aldrich
Sodium fluoride, BioXtra, ≥99%
Sigma-Aldrich
Manganese(II) chloride tetrahydrate, BioReagent, suitable for insect cell culture
Sigma-Aldrich
L-(+)-Tartaric acid, ≥99.5%
Sigma-Aldrich
Calcium chloride dihydrate, BioXtra, ≥99.0%
Sigma-Aldrich
Magnesium chloride, BioReagent, suitable for insect cell culture, ≥97.0%
Sigma-Aldrich
Manganese(II) chloride tetrahydrate, 99.99% trace metals basis
Sigma-Aldrich
Calcium chloride dihydrate, puriss. p.a., ACS reagent, reag. Ph. Eur., ≥99%
Sigma-Aldrich
(−)-Tetramisole hydrochloride, ≥99% (GC)
Sigma-Aldrich
Magnesium chloride solution, PCR Reagent, 25 mM MgCI2 solution for PCR
Sigma-Aldrich
Calcium chloride, AnhydroBeads, −10 mesh, ≥99.9% trace metals basis
Sigma-Aldrich
Magnesium chloride solution, BioUltra, for molecular biology, 2 M in H2O
Sigma-Aldrich
Sodium fluoride, 99.99% trace metals basis
Supelco
Fluoride ion solution for ISE, 0.1 M F-, analytical standard (for ion-selective electrodes)
Sigma-Aldrich
Sodium fluoride 0.5 M solution