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New cofactors and inhibitors for a DNA-cleaving DNAzyme: superoxide anion and hydrogen peroxide mediated an oxidative cleavage process.

ABSTRACT: Herein, we investigated the effects of new cofactors and inhibitors on an oxidative cleavage of DNA catalysis, known as a pistol-like DNAzyme (PLDz), to discuss its catalytic mechanism. PLDz performed its catalytic activity in the presence of ascorbic acid (AA), in which Cu²⁺ promoted, whereas Fe²⁺ significantly inhibited the catalytic function. Since Fe²⁺/AA-generated hydroxyl radicals are efficient on DNA damage, implying that oxidative cleavage of PLDz had no relation with hydroxyl radical. Subsequently, we used Fe²⁺/H₂O₂ and Cu²⁺/H₂O₂ to identify the role of hydroxyl radicals in PLDz catalysis. Data showed that PLDz lost its activity with Fe²⁺/H₂O₂, but exhibited significant cleavage with Cu²⁺/H₂O₂. Because Fe²⁺/H₂O₂ and Cu²⁺/H₂O₂ are popular reagents to generate hydroxyl radicals and the latter also produces superoxide anions, we excluded the possibility that hydroxyl radical participated in oxidative cleavage and confirmed that superoxide anion was involved in PLDz catalysis. Moreover, pyrogallol, riboflavin and hypoxanthine/xanthine oxidase with superoxide anion and hydrogen peroxide generation also induced self-cleavage of PLDz, where catalase inhibited but superoxide dismutase promoted the catalysis, suggesting that hydrogen peroxide played an essential role in PLDz catalysis. Therefore, we proposed a catalytic mechanism of PLDz in which superoxide anion and hydrogen peroxide mediated an oxidative cleavage process.

SUBMITTER: Sun Y

PROVIDER: S-EPMC5428237 | biostudies-literature | 2017 Mar

REPOSITORIES: biostudies-literature

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New cofactors and inhibitors for a DNA-cleaving DNAzyme: superoxide anion and hydrogen peroxide mediated an oxidative cleavage process.

Sun Yanhong Y Ma Rulong R Wang Shijin S Li Guiying G Sheng Yongjie Y Rui Hongyue H Zhang Jin J Xu Jiacui J Jiang Dazhi D

Scientific reports 20170323 1

Herein, we investigated the effects of new cofactors and inhibitors on an oxidative cleavage of DNA catalysis, known as a pistol-like DNAzyme (PLDz), to discuss its catalytic mechanism. PLDz performed its catalytic activity in the presence of ascorbic acid (AA), in which Cu2+ promoted, whereas Fe2+ significantly inhibited the catalytic function. Since Fe2+/AA-generated hydroxyl radicals are efficient on DNA damage, implying that oxidative cleavage of PLDz had no ...[more]

PMID: 28336968

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Project description:Skeletal muscle generation of reactive oxygen species (ROS) is increased following contractile activity and these species interact with multiple signaling pathways to mediate adaptations to contractions. The sources and time course of the increase in ROS during contractions remain undefined. Confocal microscopy with specific fluorescent probes was used to compare the activities of superoxide in mitochondria and cytosol and the hydrogen peroxide content of the cytosol in isolated single mature skeletal muscle (flexor digitorum brevis) fibers prior to, during, and after electrically stimulated contractions. Superoxide in mitochondria and cytoplasm were assessed using MitoSox red and dihydroethidium (DHE) respectively. The product of superoxide with DHE, 2-hydroxyethidium (2-HE) was acutely increased in the fiber cytosol by contractions, whereas hydroxy-MitoSox showed a slow cumulative increase. Inhibition of nitric oxide synthases increased the contraction-induced formation of hydroxy-MitoSox only with no effect on 2-HE formation. These data indicate that the acute increases in cytosolic superoxide induced by contractions are not derived from mitochondria. Data also indicate that, in muscle mitochondria, nitric oxide (NO) reduces the availability of superoxide, but no effect of NO on cytosolic superoxide availability was detected. To determine the relationship of changes in superoxide to hydrogen peroxide, an alternative specific approach was used where fibers were transduced using an adeno-associated viral vector to express the hydrogen peroxide probe, HyPer within the cytoplasmic compartment. HyPer fluorescence was significantly increased in fibers following contractions, but surprisingly followed a relatively slow time course that did not appear directly related to cytosolic superoxide. These data demonstrate for the first time temporal and site specific differences in specific ROS that occur in skeletal muscle fibers during and after contractile activity.

Dataset Information

New cofactors and inhibitors for a DNA-cleaving DNAzyme: superoxide anion and hydrogen peroxide mediated an oxidative cleavage process.

Publications

New cofactors and inhibitors for a DNA-cleaving DNAzyme: superoxide anion and hydrogen peroxide mediated an oxidative cleavage process.

Similar Datasets

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