Skip to Content
MilliporeSigma
  • Identification of the nicotinamide mononucleotide adenylyltransferase of Trypanosoma cruzi.

Identification of the nicotinamide mononucleotide adenylyltransferase of Trypanosoma cruzi.

Memorias do Instituto Oswaldo Cruz (2015-11-13)
Carlos H Niño, Nicolás Forero-Baena, Luis E Contreras, Diana Sánchez-Lancheros, Katherine Figarella, María H Ramírez
ABSTRACT

The intracellular parasite Trypanosoma cruzi is the aetiological agent of Chagas disease, a public health concern with an increasing incidence rate. This increase is due, among other reasons, to the parasite's drug resistance mechanisms, which require nicotinamide adenine dinucleotide (NAD+). Furthermore, this molecule is involved in metabolic and intracellular signalling processes necessary for the survival of T. cruzi throughout its life cycle. NAD+biosynthesis is performed by de novo and salvage pathways, which converge on the step that is catalysed by the enzyme nicotinamide mononucleotide adenylyltransferase (NMNAT) (enzyme commission number: 2.7.7.1). The identification of the NMNAT of T. cruzi is important for the development of future therapeutic strategies to treat Chagas disease. In this study, a hypothetical open reading frame (ORF) for NMNAT was identified in the genome of T. cruzi.The corresponding putative protein was analysed by simulating structural models. The ORF was amplified from genomic DNA by polymerase chain reaction and was further used for the construction of a corresponding recombinant expression vector. The expressed recombinant protein was partially purified and its activity was evaluated using enzymatic assays. These results comprise the first identification of an NMNAT in T. cruzi using bioinformatics and experimental tools and hence represent the first step to understanding NAD+ metabolism in these parasites.

MATERIALS
Product Number
Brand
Product Description

SAFC
Sodium chloride solution, 5 M
SAFC
HEPES
SAFC
HEPES
Sigma-Aldrich
Sodium hydroxide, SAJ first grade, ≥95.0%
Sigma-Aldrich
Sodium hydroxide, beads, 16-60 mesh, reagent grade, 97%
Sigma-Aldrich
Sodium hydroxide, puriss. p.a., ACS reagent, reag. Ph. Eur., K ≤0.02%, ≥98%, pellets
Sigma-Aldrich
Sodium hydroxide, reagent grade, 97%, powder
Sigma-Aldrich
Sodium hydroxide solution, purum, ≥32%
Sigma-Aldrich
Sodium hydroxide, puriss. p.a., ACS reagent, K ≤0.02%, ≥98.0% (T), pellets
Sigma-Aldrich
Imidazole, ReagentPlus®, 99%, Redi-Dri, free-flowing
Sigma-Aldrich
Imidazole, for molecular biology, ≥99% (titration), free-flowing, Redi-Dri
Sigma-Aldrich
Sodium hydroxide, anhydrous, free-flowing, Redi-Dri, reagent grade, ≥98%, pellets
Sigma-Aldrich
Imidazole, ACS reagent, ≥99% (titration)
Sigma-Aldrich
Sodium hydroxide solution, BioUltra, for molecular biology, 10 M in H2O
Sigma-Aldrich
Sodium chloride, BioUltra, for molecular biology, ≥99.5% (AT)
Sigma-Aldrich
Sodium chloride, random crystals, optical grade, 99.9% trace metals basis
Sigma-Aldrich
Sodium chloride-35Cl, 99 atom % 35Cl
Sigma-Aldrich
3-Ethyl-2,4-pentanedione, mixture of tautomers, 98%
Sigma-Aldrich
β-Nicotinamide mononucleotide, ≥95% (HPLC)
Sigma-Aldrich
Sodium hydroxide, JIS special grade, ≥96.0%
Sigma-Aldrich
Sodium chloride, SAJ first grade, ≥99.0%
Sigma-Aldrich
Sodium chloride, JIS special grade, ≥99.5%
Sigma-Aldrich
Sodium hydroxide, pellets, semiconductor grade, 99.99% trace metals basis
Sigma-Aldrich
Sodium hydroxide solution, 5.0 M
Sigma-Aldrich
Sodium hydroxide, BioUltra, for luminescence, ≥98.0% (T), pellets
Sigma-Aldrich
Sodium hydroxide, puriss., meets analytical specification of Ph. Eur., BP, NF, E524, 98-100.5%, pellets
Sigma-Aldrich
Sodium chloride solution, BioUltra, for molecular biology, ~5 M in H2O
Sigma-Aldrich
Sodium chloride solution, 5 M in H2O, BioReagent, for molecular biology, suitable for cell culture
Sigma-Aldrich
Sodium chloride, for molecular biology, DNase, RNase, and protease, none detected, ≥99% (titration)
Sigma-Aldrich
Potassium phosphate tribasic, reagent grade, ≥98%