• The Korean Journal of Physiology and Pharmacology
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• v.20 no.5
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• pp.449-457
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• 2016

N-acetyl-L-cysteine (NAC) and cysteine have been implicated in a number of human neutrophils' functional responses. However, though $Ca^{2+}$ signaling is one of the key signalings contributing to the functional responses of human neutrophils, effects of NAC and cysteine on intracellular calcium concentration ($[Ca^{2+}]_i$) in human neutrophils have not been investigated yet. Thus, this study was carried out with an objective to investigate the effects of NAC and cysteine on $[Ca^{2+}]_i$ in human neutrophils. We observed that NAC ($1{\mu}M{\sim}1mM$) and cysteine ($10{\mu}M{\sim}1mM$) increased $[Ca^{2+}]_i$ in human neutrophils in a concentration-dependent manner. In NAC pre-supplmented buffer, an additive effect on N-formyl-methionine-leucine-phenylalanine (fMLP)-induced increase in $[Ca^{2+}]_i$ in human neutrophils was observed. In $Ca^{2+}$-free buffer, NAC- and cysteine-induced $[Ca^{2+}]_i$ increase in human neutrophils completely disappeared, suggesting that NAC- and cysteine-mediated increase in $[Ca^{2+}]_i$ in human neutrophils occur through $Ca^{2+}$ influx. NAC- and cysteine-induced $[Ca^{2+}]_i$ increase was effectively inhibited by calcium channel inhibitors SKF96365 ($10{\mu}m$) and ruthenium red ($20{\mu}m$). In $Na^+$-free HEPES, both NAC and cysteine induced a marked increase in $[Ca^{2+}]_i$ in human neutrophils, arguing against the possibility that $Na^+$-dependent intracellular uptake of NAC and cysteine is necessary for their $[Ca^{2+}]_i$ increasing activity. Our results show that NAC and cysteine induce $[Ca^{2+}]_i$ increase through $Ca^{2+}$ influx in human neutrophils via SKF96365- and ruthenium red-dependent way.

• Molecules and Cells
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• v.26 no.3
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• pp.228-235
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• 2008

Thiol-dependent redox systems are involved in regulation of diverse biological processes, such as response to stress, signal transduction, and protein folding. The thiol-based redox control is provided by mechanistically similar, but structurally distinct families of enzymes known as thiol oxidoreductases. Many such enzymes have been characterized, but identities and functions of the entire sets of thiol oxidoreductases in organisms are not known. Extreme sequence and structural divergence makes identification of these proteins difficult. Thiol oxidoreductases contain a redox-active cysteine residue, or its functional analog selenocysteine, in their active sites. Here, we describe computational methods for in silico prediction of thiol oxidoreductases in nucleotide and protein sequence databases and identification of their redox-active cysteines. We discuss different functional categories of cysteine residues, describe methods for discrimination between catalytic and noncatalytic and between redox and non-redox cysteine residues and highlight unique properties of the redox-active cysteines based on evolutionary conservation, secondary and three-dimensional structures, and sporadic replacement of cysteines with catalytically superior selenocysteine residues.

• Journal of Applied Biological Chemistry
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• v.53 no.4
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• pp.195-201
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• 2010

A gene encoding an alanine racemase in B. pseudomycoides was cloned and one (Cys316) or both of two cysteines (Cys316 and Cys365) was (were) substituted with alanine. The cysteine (-) alanine racemases were expressed in E. coli BL21 (DE3) using a pET-21 vector. The expressed enzymes were purified through affinity chromatography using 6xHis ligand. The purified enzymes all showed major one bands by SDS-PAGE analysis, corresponding to 46 kDa. The cysteine (-) alanine racemases as well as the wild type enzyme showed alanine racemase activities, indicating that the enzyme is an alanine racemase and the cysteines in the enzyme may not be involved in the catalysis and/or substrate binding. Thermal stabilities of Cys (-) alanine racemases decreased considerably and half-lives were 26 (wild type), 21 (C316A) and 18 min (C316-365A), respectively at $60^{\circ}C$ pH 8.0, suggesting that cysteine is considerably contributive to the thermal stability of the alanine racemase.

• Journal of Applied Biological Chemistry
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• v.53 no.3
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• pp.132-138
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• 2010

A gene encoding a putative alanine racemase in B. pseudomycoides was cloned and expressed in Escherichia coli BL21(DE3) using a pET-21 vector harbouring 6xHistidine tag. Affinity purification of the recombinant alanine racemase with a nickel resin resulted in one band by SDS-PAGE analysis. The purified enzyme showed a molecular weight of 46 kDa. The enzyme was the most active toward L-alanine and secondly D-alanine, implying that the enzyme is an alanine racemase. D-cysteine significantly inhibited the enzyme activity and also L-cysteine to a lesser extent. The enzyme was considerably activated by addition of pyridoxal-5'-phosphate (PLP), showing that 73% increase in activity was observed at 0.3 mM, compared to control. The enzyme was the most active at pH 9.0 and more stable at alkaline pHs than acidic pHs.

• Parasites, Hosts and Diseases
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• v.43 no.4 s.136
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• pp.157-160
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• 2005

A 29kDa cysteine protease of Taenia solium metacestodes was purified by Mono Q anion-exchanger and Superose 6 HR gel filtration chromatography. The enzyme was effectively inhibited by cysteine protease inhibitors, such as iodoacetic acid (IAA) and trans-epoxy-succinyl-L-leucyl-amido (4-guanidino) butane (E-64) while inhibitors acting on serine- or metallo-proteases did not affect the enzyme activity. The purified enzyme degraded human immunoglobulin G (IgG), collagen and bovine serum albumin (BSA), but human IgG was more susceptible for proteolysis by the enzyme. To define the precise biological roles of the enzyme, more detailed biochemical and functional studies would be required.

• Food Science and Preservation
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• v.20 no.2
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• pp.173-181
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• 2013

This study investigated that anti-browning effects of Atractylodis Rhizoma Alba extract and L-cysteine combination. Mushrooms were dipped in solutions (0.1% Atractylodis Rhizoma Alba extract containing 0.05% L-cysteine) for 3 min. The dipped mushrooms were packaged in a polystyrene (PS) tray and wrapped with a polyvinyl chloride (PVC) film, and stored for 14 days at $10^{\circ}C$. The browning inhibition activity (Hunter L, a, b color scale and tyrosinase inhibition activity) and quality changes (weight loss rate, gas composition, firmness and sensory evaluation) were analyzed during storage period. After 14 days, the Hunter L and ${\Delta}E$ value of mushrooms treated in 0.1% Atractylodis Rhizoma Alba extract containing 0.05% L-cysteine were 87.24 and 5.56, respectively. The mushrooms treated with 0.1% Atractylodis Rhizoma Alba extract containing 0.05% L-cysteine also showed higher firmness (13.31 N) and smaller weight loss rate (2.87%) than the untreated mushroom (11.42 N, 3.04%) on storage day 14. During storage period, the sensory evaluation showed that overall acceptability of mushrooms treated with 0.1% Atractylodis Rhizoma Alba extract containing 0.05% L-cysteine were higher than those of the untreated mushrooms, except those that were stored for five days. Overall, the mushrooms treated with 0.1% Atractylodis Rhizoma Alba extract containing 0.05% L-cysteine had a higher tyrosinase inhibition activity than the untreated mushrooms during storage period. This study suggests that the browning of the mushrooms treated with 0.1% Atractylodis Rhizoma Alba extract containing 0.05% L-cysteine solution were inhibited, and the that their shelf life was extended.

• Journal of Plant Biotechnology
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• v.1 no.1
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• pp.13-19
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• 1999

The visco-elastic properties of gluten are major determinants of the processing properties of doughs. These visco-elastic properties are strongly influenced by the ratio of monomeric and polymeric proteins and the size distribution of the polymeric proteins, which make up the gluten fraction of the dough. Recent studies have revealed that other features, such as the number of the cysteine residues of the HMW-GS, also play an important role in determining the functional characteristics. To modify the processing properties at molecular level, the relationship between the structure of molecules and dough properties has to be understood. In order to explore the relationships between individual proteins and dough properties, we have developed procedures for incorporating bacterially expressed proteins into doughs, and measuring their functional properties in small-scale equipment. A major problem in investigating the structure/function relationships of individual seed storage proteins is to obtain sufficient amounts of pure polypeptides from the complex families of proteins expressed in the endosperm. Therefore, we have established a simplified model system in which we produce specific protein genes through bacterial expression and test their functional properties in smallscale apparatus after incorporation into base flour. An S poor protein gene has been chosen as a template gene. This template gene has been modified using standard recombinant DNA techniques in order to test the effects of varying the number and position of cysteine residues, and the size of the protein. Doughs have been mixed in small scale apparatus and characterized with respect to their polymeric composition and their functional properties, including dough mixing, extensibility and small scale bating. We conclude that dough characteristics can be manipulated in a predictable manner by altering the cysteine residues and the size of high molecular weight glutenins.

• Asian-Australasian Journal of Animal Sciences
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• v.30 no.5
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• pp.728-735
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• 2017

Objective: This study was performed to reveal the molecular structure and expression patterns of horse glutamate-cysteine ligase catalytic subunit (GCLC) and glutamate-cysteine ligase modifier subunit (GCLM) genes whose products form glutamate cysteine ligase, which were identified as differentially expressed genes in the previous study. Methods: We performed bioinformatics analyses, and gene expression assay with quantitative polymerase chain reaction (qPCR) for horse GCLC and GCLM genes in muscle and blood leukocytes of Thoroughbred horses Results: Expression of GCLC showed the same pattern in both blood and muscle tissues after exercise. Expression of GCLC increased in the muscle and blood of Thoroughbreds, suggesting a tissue-specific regulatory mechanism for the expression of GCLC. In addition, expression of the GCLM gene increased after exercise in both the blood and muscle of Thoroughbreds. Conclusion: We established the expression patterns of GCLC and GCLM in the skeletal muscle and blood of Thoroughbred horses in response to exercise. Further study is now warranted to uncover the functional importance of these genes in exercise and recovery in racehorses.

• Preventive Nutrition and Food Science
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• v.19 no.3
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• pp.220-226
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• 2014

In the present report, the effects of blanching, steaming, and high temperature/high pressure processing (HTHP) on the amino acid contents of commonly consumed Korean root vegetables, leaf vegetables, and pulses were evaluated using an Automatic Amino Acid Analyzer. The total amino acid content of the samples tested was between 3.38 g/100 g dry weight (DW) and 21.32 g/100 g DW in raw vegetables and between 29.36 g/100 g DW and 30.55 g/100 g DW in raw pulses. With HTHP, we observed significant decreases in the lysine and arginine contents of vegetables and the lysine, arginine, and cysteine contents of pulses. Moreover, the amino acid contents of blanched vegetables and steamed pulses were more similar than the amino acid contents of the HTHP vegetables and HTHP pulses. Interestingly, lysine, arginine, and cysteine were more sensitive to HTHP than the other amino acids. Partial Least Squares-Discriminate Analyses were also performed to discriminate the clusters and patterns of amino acids.

• BMB Reports
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• v.43 no.1
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• pp.23-28
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• 2010

Binary toxin consisting of BinA and BinB from Bacillus sphaericus is toxic to mosquito larvae. BinB is responsible for specific binding to the larval gut cell membrane while BinA is crucial for toxicity. To investigate functional role of cysteine in BinB, three cysteine residues at positions 67, 161, and 241 were replaced by alanine or serine. Mutations at these positions did not affect protein production and overall structure of BinB. These cysteine residues are not involved in disulfide bond formation between BinB molecules. Mosquito-larvicidal assays revealed that C67 and C161 are essential for toxicity, whereas C241 is not. Mutations at C67 and C161 resulted in weaker BinA-BinB interaction. The loss of toxicity may be due to the reduction of interactions between BinA and BinB or BinB and its receptor. C67 and C161 could also play a part during conformational changes or internalization of the binary toxin into the target cell.