Nitric oxide cytotoxicity and functions of iron-sulfur enzymes in Escherichia coli

Iron-sulfur proteins are ubiquitous in biological processes. Here we report that NO, a physiological free radical, can effectively inhibit cell growth of Escherichia coli in minimal medium under anaerobic growth conditions. Fractionation of the cell extracts obtained from NO-exposed cells shows a broad distribution of the protein-bound dinitrosyl-iron complexes (DNICs) formed by NO. On the other hand, the cell growth of E. coli can be restored when NO-exposed cells are either supplemented with the branched-chain amino acids (BCAAs) anaerobically or returned to aerobic growth conditions. It turns out that dihydroxyacid dehydratase (IlvD), an iron-sulfur enzyme essential for the BCAAs biosynthesis, is completely inactivated by NO along with formation of the IlvD-bound DNICs. Nevertheless, the IlvD-DNICs, together with other protein-bound DNICs, are sufficiently repaired under aerobic growth conditions without new proteins synthesis. It is proposed that cellular deficiency to repair the NO-modified iron-sulfur proteins may directly contribute to the NO-induced bacteriostasis under anaerobic conditions. We further identify a new iron-sulfur enzyme as a potential target of NO, a DNA damage-inducible helicase DinG, which is a member of the DNA helicase superfamily II involved in DNA replication and repair. We find that E. coli DinG contains a redox-active [4Fe-4S] cluster with a midpoint redox potential (Em) of -390 ± 23 mV (pH 8.0). An oxidized [4Fe-4S] cluster in DinG is required for its helicase activity and reduction of the [4Fe-4S] cluster can reversibly switch off its helicase activity. While DinG is resistant to 100-fold excess of hydrogen peroxide, it can be readily inactivated by NO, implying that inactivation of DinG by NO could impact the DNA repair and result in genomic instability. The NO-modified DinG can be fully reactivated by reassembly of a new [4Fe-4S] cluster, indicating that DinG is the primary target of NO exposure. The results from this study suggest that NO can effectively modify the iron-sulfur cluster in proteins, thus contributing to the NO-mediated bacteriostasis and genomic instability

CORRELATION BETWEEN PERCEIVED SOCIAL SUPPORT AND LONELINESS AMONG CHINESE ADOLESCENTS: MEDIATING EFFECTS OF PSYCHOLOGICAL CAPITAL

Background: Adolescence is a period characterized by high frequency of mental health problems. Loneliness, in particular, is a common psychological problem. This study aims to explore the effects of psychological capital on adolescents’ loneliness and the mediating effects of perceived social support. Subjects and methods: A total of 694 adolescents from six regions in Hunan Province, China were selected. The survey was conducted using the Perceived Social Support Scale, Loneliness Scale, and Psychological Capital Questionnaire. The study analyzed the correlations between perceived social support, psychological capital, and loneliness among adolescents and the mediating role of psychological capital in the relationship between perceived social support and loneliness. Results: Significant correlations were observed between adolescents’ total scores in perceived social support, psychological capital, and loneliness and between each dimension. The following observations were made: a significant negative correlation between perceived social support and loneliness (r= 0.440, P<0.01), a significant positive correlation between perceived social support and psychological capital (r=0.493, P<0.01), and a significant negative correlation between psychological capital and loneliness (r=-0.303, P<0.01). Psychological capital plays a statistically significant mediating role on the relationship between perceived social support and loneliness (P<0.01). Conclusions: Perceived social support and psychological capital can reduce adolescents’ loneliness and perceived social support can alleviate loneliness by enhancing psychological capital

Experimental Investigation into the Effect of Pyrolysis on Chemical Forms of Heavy Metals in Sewage Sludge Biochar (SSB), with Brief Ecological Risk Assessment

publishedVersio

Stimulation of Escherichia coli DNA damage inducible DNA helicase DinG by the single-stranded DNA binding protein SSB

Escherichia coli DNA damage inducible protein DinG is a superfamily II DNA helicase and is closely related to human DNA helicase XPD. Here, we report that E. coli single-stranded DNA binding protein (SSB) is able to form a stable protein complex with DinG and to stimulate the DinG DNA helicase activity. An SSB mutant that retains the single-stranded DNA binding activity but fails to form a protein complex with DinG becomes a potent inhibitor for the DinG DNA helicase, suggesting that E. coli wild-type SSB stimulates the DinG DNA helicase via specific protein-protein interaction. Structured summary of protein interactions: SSB and SSB bind by molecular sieving (View interaction) DinG and SSB bind by molecular sieving (View interaction) DinG and SSB bind by cosedimentation in solution (View interaction) © 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved

Nitric oxide-induced bacteriostasis and modification of iron-sulphur proteins in Escherichia coli

The nitric oxide (NO) cytotoxicity has been well documented in bacteria and mammalian cells. However, the underlying mechanism is still not fully understood. Here we report that transient NO exposure effectively inhibits cell growth of Escherichia coli in minimal medium under anaerobic growth conditions and that cell growth is restored when the NO-exposed cells are either supplemented with the branched-chain amino acids (BCAA) anaerobically or returned to aerobic growth conditions. The enzyme activity measurements show that dihydroxyacid dehydratase (IlvD), an iron-sulphur enzyme essential for the BCAA biosynthesis, is completely inactivated in cells by NO with the concomitant formation of the IlvD-bound dinitrosyl iron complex (DNIC). Fractionation of the cell extracts prepared from the NO-exposed cells reveals that a large number of different protein-bound DNICs are formed by NO. While the IlvD-bound DNIC and other protein-bound DNICs are stable in cells under anaerobic growth conditions, they are efficiently repaired under aerobic growth conditions even without new protein synthesis. Additional studies indicate that l-cysteine may have an important role in repairing the NO-modified iron-sulphur proteins in aerobically growing E. coli cells. The results suggest that cellular deficiency to repair the NO-modified iron-sulphur proteins may directly contribute to the NO-induced bacteriostasis under anaerobic conditions. © 2008 The Authors

Reactivity of nitric oxide with the [4Fe-4S] cluster of dihydroxyacid dehydratase from Escherichia coli

Although the NO (nitric oxide)-mediated modification of iron-sulfur proteins has been well-documented in bacteria and mammalian cells, specific reactivity of NO with iron-sulfur proteins still remains elusive. In the present study, we report the first kinetic characterization of the reaction between NO and iron-sulfur clusters in protein using the Escherichia coli IlvD (dihydroxyacid dehydratase) [4Fe-4S] cluster as an example. Combining a sensitive NO electrode with EPR (electron paramagnetic resonance) spectroscopy and an enzyme activity assay, we demonstrate that NO is rapidly consumed by the IlvD [4Fe-4S] cluster with the concomitant formation of the IlvD-bound DNIC (dinitrosyl-iron complex) and inactivation of the enzyme activity under anaerobic conditions. The rate constant for the initial reaction betweenNOand the IlvD [4Fe-4S] cluster is estimated to be (7.0±2.0)×106 M-2 · s-1 at 25°C, which is approx. 2-3 times faster than that of the NO autoxidation by O2 in aqueous solution. Addition of GSH failed to prevent the NOmediated modification of the IlvD [4Fe-4S] cluster regardless of the presence of O2 in the medium, further suggesting that NO is more reactivewith the IlvD [4Fe-4S] cluster than with GSH or O2. Purified aconitase B [4Fe-4S] cluster from E. coli has an almost identical NO reactivity as the IlvD [4Fe-4S] cluster. However, the reaction between NO and the endonuclease III [4Fe-4S] cluster is relatively slow, apparently because the [4Fe-4S] cluster in endonuclease III is less accessible to solvent than those in IlvD and aconitase B. When E. coli cells containing recombinant IlvD, aconitase B or endonuclease III are exposed to NO using the Silastic tubing NO delivery system under aerobic and anaerobic conditions, the [4Fe-4S] clusters in IlvD and aconitase B, but not in endonuclease III, are efficiently modified forming the proteinbound DNICs, confirming that NO has a higher reactivity with the [4Fe-4S] clusters in IlvD and aconitase B than with O2 or GSH. The results suggest that the iron-sulfur clusters in proteins such as IlvD and aconitase B may constitute the primary targets of the NO cytotoxicity under both aerobic and anaerobic conditions. © The Authors Journal compilation © 2009 Biochemical Society

Nitrogen-doped graphene-ionic liquid-glassy carbon microsphere paste electrode for ultra-sensitive determination of quercetin

The analysis of quercetin (Qu) is of great significance owing to its multiple biomedical effects. In this work, a nitrogendoped graphene-ionic liquid-glassy carbon microsphere paste electrode (N-GE/GCILE) was constructed for the determination of Qu. Cyclic voltammetry (CV) and square wave voltammetry (SWV) were employed to investigate the electrochemical behavior of Qu. In comparison with unmodified glassy carbon microsphere paste electrode, the modified electrode exhibited better electrocatalytic activity towards Qu. The influencing conditions on sensitivity such as the amount of modifier, accumulation potential and time, and electrolyte pH value were respectively discussed. Under the optimized conditions, two linear ranges of 0.002- 0.1 μM and 0.1-10 μM were obtained, with a detection limit of 1 nM (S/N=3). The method was applied in Qu determination in blueberry juice with the recoveries of 102.5-105.0 %

Monitoring of atopic dermatitis using leaky coaxial cable

In our daily life, inadvertent scratching may increase the severity of skin diseases (such as atopic dermatitis, etc.). However, people rarely pay attention to this matter, so the known measurement behavior of the movement is also very little. Nevertheless, the behavior and frequency of scratching represent the degree of itching, and the analysis of scratching frequency is helpful to the doctor's clinical dosage. In this paper, a novel system is proposed to monitor the scratching motion of a sleeping human body at night. The core device of the system are just a Leaky coaxial cable (LCX) and a router. Commonly, LCX is used in the blind field or semi blind field in wireless communication. The new idea is that the leaky cable is placed on the bed, then the state information of physical layer of wireless communication channels is acquired to identify the scratching motion and other small body movements in the human sleep process. The results show that it can be used to detect the movement and its duration. Channel state information (CSI) packet is collected by card installed in the computer based on the 802.11n protocol. The characterization of the scratch motion in the collected channel state information is unique, so it can be distinguished from the wireless channel amplitude variation trend