Limits...
Identification of the nicotinamide mononucleotide adenylyltransferase of Trypanosoma cruzi.

Niño CH, Forero-Baena N, Contreras LE, Sánchez-Lancheros D, Figarella K, Ramírez MH - Mem. Inst. Oswaldo Cruz (2015)

Bottom Line: 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.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.

View Article: PubMed Central - PubMed

Affiliation: Laboratorio de Investigaciones Básicas en Bioquímica, Facultad de Ciencias, Universidad Nacional de Colombia, Bogotá, Colombia.

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.

Show MeSH

Related in: MedlinePlus

Secondary structure prediction for a nicotinamide mononucleotideadenylyltransferase of Trypanosoma cruzi candidate
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4660618&req=5

f07: Secondary structure prediction for a nicotinamide mononucleotideadenylyltransferase of Trypanosoma cruzi candidate

Mentions: Bioinformatics analysis of the putative TcNMNAT - Using the sequencefrom the genome of CL Brener Esmeraldo-like (TcCLB.507047.170), a structural analysis ofthe putative TcNMNAT was performed. The primary structure showed that the putativeprotein is 289 aa and has a molecular mass of 32,031.4 Da. The modelling of thesecondary structure pattern was performed using three different algorithms (Table II). The GORIV algorithm (precision ~64%) isbased on the propensity of each aa and its neighbours to form a determined secondarystructure using a dynamic window of a determined length. The third-generation algorithmPDH (precision >70%) is based on adaptive neural networks and the PREDATOR algorithm(precision ~68-75%) is based on the recognition of aa that are potentially linked byhydrogen bonds. All three bioinformatics methods showed similar results, with a mean of43.90% residues located in α-helices, 12.57% in β-sheets and 44.52% in randomregions.


Identification of the nicotinamide mononucleotide adenylyltransferase of Trypanosoma cruzi.

Niño CH, Forero-Baena N, Contreras LE, Sánchez-Lancheros D, Figarella K, Ramírez MH - Mem. Inst. Oswaldo Cruz (2015)

Secondary structure prediction for a nicotinamide mononucleotideadenylyltransferase of Trypanosoma cruzi candidate
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4660618&req=5

f07: Secondary structure prediction for a nicotinamide mononucleotideadenylyltransferase of Trypanosoma cruzi candidate
Mentions: Bioinformatics analysis of the putative TcNMNAT - Using the sequencefrom the genome of CL Brener Esmeraldo-like (TcCLB.507047.170), a structural analysis ofthe putative TcNMNAT was performed. The primary structure showed that the putativeprotein is 289 aa and has a molecular mass of 32,031.4 Da. The modelling of thesecondary structure pattern was performed using three different algorithms (Table II). The GORIV algorithm (precision ~64%) isbased on the propensity of each aa and its neighbours to form a determined secondarystructure using a dynamic window of a determined length. The third-generation algorithmPDH (precision >70%) is based on adaptive neural networks and the PREDATOR algorithm(precision ~68-75%) is based on the recognition of aa that are potentially linked byhydrogen bonds. All three bioinformatics methods showed similar results, with a mean of43.90% residues located in α-helices, 12.57% in β-sheets and 44.52% in randomregions.

Bottom Line: 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.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.

View Article: PubMed Central - PubMed

Affiliation: Laboratorio de Investigaciones Básicas en Bioquímica, Facultad de Ciencias, Universidad Nacional de Colombia, Bogotá, Colombia.

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.

Show MeSH
Related in: MedlinePlus