Dataset Information

Mouse Nudt13 is a Mitochondrial Nudix Hydrolase with NAD(P)H Pyrophosphohydrolase Activity.

ABSTRACT: The mammalian NUDT13 protein possesses a sequence motif characteristic of the NADH pyrophosphohydrolase subfamily of Nudix hydrolases. Due to the persistent insolubility of the recombinant product expressed in Escherichia coli, active mouse Nudt13 was expressed in insect cells from a baculovirus vector as a histidine-tagged recombinant protein. In vitro, it efficiently hydrolysed NADH to NMNH and AMP and NADPH to NMNH and 2',5'-ADP and had a marked preference for the reduced pyridine nucleotides. Much lower activity was obtained with other nucleotide substrates tested. K m and k cat values for NADH were 0.34 mM and 7 s-1 respectively. Expression of Nudt13 as an N-terminal fusion to green fluorescent protein revealed that it was targeted exclusively to mitochondria by the N-terminal targeting peptide, suggesting that Nudt13 may act to regulate the concentration of mitochondrial reduced pyridine nucleotide cofactors and the NAD(P)+/NAD(P)H ratio in this organelle and elsewhere. Future studies of the enzymology of pyridine nucleotide metabolism in relation to energy homeostasis, redox control, free radical production and cellular integrity should consider the possible regulatory role of Nudt13.

SUBMITTER: Abdelraheim SR

PROVIDER: S-EPMC5626787 | biostudies-literature | 2017 Oct

REPOSITORIES: biostudies-literature

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Mouse Nudt13 is a Mitochondrial Nudix Hydrolase with NAD(P)H Pyrophosphohydrolase Activity.

Abdelraheim Salama R SR Spiller David G DG McLennan Alexander G AG

The protein journal 20171001 5

The mammalian NUDT13 protein possesses a sequence motif characteristic of the NADH pyrophosphohydrolase subfamily of Nudix hydrolases. Due to the persistent insolubility of the recombinant product expressed in Escherichia coli, active mouse Nudt13 was expressed in insect cells from a baculovirus vector as a histidine-tagged recombinant protein. In vitro, it efficiently hydrolysed NADH to NMNH and AMP and NADPH to NMNH and 2',5'-ADP and had a marked preference for the reduced pyridine nucleotides ...[more]

PMID: 28755312

Similar Datasets

Project description:The PA0336 protein from Pseudomonas aeruginosa belongs to the family of widely distributed Nudix pyrophosphohydrolases which catalyze the hydrolysis of pyrophosphate bonds in a variety of nucleoside diphosphate derivatives. The amino acid sequence of the PA0336 protein is highly similar to that of the RppH Nudix RNA pyrophosphohydrolase from E. coli which removes pyrophosphate from 5’-end of triphosphorylated RNA transcripts. Trans-complementation experiments showed that the P. aeruginosa enzyme can functionally substitute for RppH in E. coli cells indicating that, similarly to RppH, the Pseudomonas hydrolase mediates RNA turnover in vivo. In order to elucidate the biological significance of the PA0336 protein in Pseudomonas cells, a PA0336 mutant strain was constructed. The mutated strain considerably increased level of the virulence factor pyocyanin compared to wild type, suggesting that PA0336 could be involved in down-regulation of P. aeruginosa pathogenicity. This phenotype was reversed by complementation with the wild type, but not catalytically inactive PA0336, indicating that the catalytic activity was indispensable for its biological function. To study the role of PA0336 further, transcriptomes of the PA0336 mutant and the wild type strain were compared using RNA sequencing. The cellular level of a number of transcripts was affected by the lack of PA0336. We focused our attention on pathogenesis-related genes. Up-regulated in the PA0336 mutant were transcripts coding for, i. a., proteins involved in the regulation and/or production of pyocyanin, biofim-associated alginates and exotoxins. The results from the global analysis were verified by determining the cellular level of chosen transcripts by quantitative RT-PCR method. Pathogenesis tests in Caenorhabditis elegans showed that the PA0336 mutant of P. aeruginosa was significantly more virulent than the parental strain, confirming further that the P. aeruginosa RNA pyrophosphohydrolase PA0336 modulates bacterial pathogenesis by down-regulating production of virulence factors.

Dataset Information

Mouse Nudt13 is a Mitochondrial Nudix Hydrolase with NAD(P)H Pyrophosphohydrolase Activity.

Publications

Mouse Nudt13 is a Mitochondrial Nudix Hydrolase with NAD(P)H Pyrophosphohydrolase Activity.

Similar Datasets

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