• BMB Reports
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• v.43 no.3
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• pp.176-181
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• 2010

The primary sigma factor ($\sigma^{A}$) of Staphylococcus aureus, a potential drug target, was little investigated at the structural level. Using an N-terminal histidine-tagged $\sigma^{A}$ (His-$\sigma^{A}$), here we have demonstrated that it exits as a monomer in solution, possesses multiple domains, harbors primarily $\alpha$-helix and efficiently binds to a S. aureus promoter DNA in the presence of core RNA polymerase. While both N- and C-terminal ends of His-$\sigma^{A}$ are flexible in nature, two Trp residues in its DNA binding region are buried. Upon increasing the incubation temperature from 25$^{\circ}$ to 40$^{\circ}C$, $\sim$60% of the input His-$\sigma^{A}$ was cleaved by thermolysin. Aggregation of His-$\sigma^{A}$ was also initiated rapidly at 45$^{\circ}C$. From the equilibrium unfolding experiment, the Gibbs free energy of stabilization of His-$\sigma^{A}$ was estimated to be +0.70 kcal $mol^{-1}$. The data together suggest that primary sigma factor of S. aureus is an unstable protein. Core RNA polymerase however stabilized $\sigma^{A}$ appreciably.

• Journal of Integrative Natural Science
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• v.4 no.4
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• pp.273-277
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• 2011

It's a threat for the public health that H1N1 (Influenza virus A) causes disease and transmits among humans. WHO (world health organization) declared that the infections caused by the new strain had reached pandemic proportions. The approved neuraminidase inhibitors (Zanamivir and Oseltamivir) and related investigative drug (BCX-1812) are potent, specific inhibitors of influenza A and B viruses. These drugs are highly effective to prevent influenza A and B infections. Early therapeutic use reduces illness duration and respiratory complications. Recently, we found one of the potent inhibitor of erystagallin A ($IC_{50}$ of 2.04 ${\mu}M$) for neuraminidase target, this inhibitor shows most similar structure to its natural substrate, sialic acid. Therefore, we chose 1l7f to get the receptor structure for docking study among many crystal structures. A docking study has been performed in Surflex-Dock module in SYBYL 8.1. In the present study, we attempt to compare the docking studies of pterocarpin and erystagallin A with neuraminidase receptor structure. In the previous report, the methoxy group of pterocarpin had H-bonding with Arg residues. The present docking results for erystagallin A showed the backbone of hydroxyl group shows significant H-bonding interactions with Arg152 and Arg292. The results showed that erystagallin A interacts more favorably with distinctive binding site rather than original active site. Therefore, we tried to reveal plausible binding mode and important amino acid for this inhibitor using docking and site id search calculations of Sybyl. The results obtained from this work may be utilized to design novel inhibitors for neuraminidase.

• Genomics & Informatics
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• v.12 no.4
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• pp.283-288
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• 2014

Among all serious diseases globally, diabetes (type 1 and type 2) still poses a major challenge to the world population. Several target proteins have been identified, and the etiology causing diabetes has been reasonably well studied. But, there is still a gap in deciding on the choice of a drug, especially when the target is mutated. Mutations in the KCNJ11 gene, encoding the kir6.2 channel, are reported to be associated with congenital hyperinsulinism, having a major impact in causing type 1 diabetes, and due to the lack of its 3D structure, an attempt has been made to predict the structure of kir6.2, applying fold recognition methods. The current work is intended to investigate the affinity of four phytochemicals namely, curcumin (Curcuma longa), genistein (Genista tinctoria), piperine (Piper nigrum), and pterostilbene (Vitis vinifera) in a normal as well as in a mutant kir6.2 model by adopting a molecular docking methodology. The phytochemicals were docked in both wild and mutated kir6.2 models in two rounds: blind docking followed by ATP-binding pocket-specific docking. From the binding pockets, the common interacting amino acid residues participating strongly within the binding pocket were identified and compared. From the study, we conclude that these phytochemicals have strong affinity in both the normal and mutant kir6.2 model. This work would be helpful for further study of the phytochemicals above for the treatment of type 1 diabetes by targeting the kir6.2 channel.

• Journal of Ginseng Research
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• v.32 no.2
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• pp.99-106
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• 2008

A ligand - whether an endogenous hormone, neurotransmitter, exogenous toxin or synthetic drug - binds to plasma membrane proteins (e.g., ion channels, receptors or other functional proteins) to exert its physiological or pharmacological effects. Ligands can also have functional groups, showing stereospecificity for interaction sites on their counterpart plasma membrane proteins. Previous reports have shown that the ginsenoside Rg$_3$, a bioactive ginsenoside, meets these criteria in that: 1) an aliphatic side chain of $Rg_3$ plays a role as a functional group, 2) Rg$_3$ regulates voltage- and ligand-gated ion channels in a stereospecific manner with respect to carbon-20, and 3) $Rg_3$ regulates subsets of ligand-gated and voltage-gated ion channels through specific interactions with identified amino acid residues inside the channel pore, in the outer pore entryway, or in toxin binding sites. Rg$_3$, therefore, could be a candidate for a novel ginseng-derived glycosidic ligand regulating ion channels and receptors. This review will examine how Rg$_3$ regulates voltage-gated and ligand-gated ion channels through interactions with its target proteins in the plasma membrane. Hopefully, this review will advance understanding of ginseng pharmacology at the cellular and molecular levels.

• Journal of Pharmacopuncture
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• v.23 no.2
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• pp.71-78
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• 2020

Objectives: This study was conducted to development of hazardous materials management standards for the decoction type of personalized herbal medicines (PHMs). Methods: This study was conducted in two stages. We searched documents about criteria to use words such as 'Herb', 'Herbal medicine', and 'Botanical Drug' and summarized the results. We organized the committee consisted of seven experts, and held two meetings to reach an agreement on hazardous management standards of the decoction type of PHMs. Results: The seven documents were presented in the literature review and six items related to hazardous management standards of decoction were identified. The second expert meeting brought that a total of six items, including heavy metal, pesticide residues, sulfur dioxide, benzopyrene, mycotoxin, and micro-organism limits, were selected for safety management of decoction type of PHMs. Also, the criteria and test methods for each standard were suggested for monitoring the decoction type of PHMs. Conclusion: The study suggested hazardous material management standards and criteria for the decoction types of PHMs. In the future, it would be necessary to conduct a pilot test to ensure the validity and credibility of the safety management standard and criteria. Furthermore, the government level safety management system should be introduced to verify the safety of decoction medicines.

• Genomics & Informatics
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• v.14 no.2
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• pp.53-61
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• 2016

Toxoplasma gondii is an intracellular Apicomplexan parasite and a causative agent of toxoplasmosis in human. It causes encephalitis, uveitis, chorioretinitis, and congenital infection. T. gondii invades the host cell by forming a moving junction (MJ) complex. This complex formation is initiated by intermolecular interactions between the two secretory parasitic proteins-namely, apical membrane antigen 1 (AMA1) and rhoptry neck protein 2 (RON2) and is critically essential for the host invasion process. By this study, we propose two potential leads, NSC95522 and NSC179676 that can efficiently target the AMA1 hydrophobic cleft, which is a hotspot for targeting MJ complex formation. The proposed leads are the result of an exhaustive conformational search-based virtual screen with multilevel precision scoring of the docking affinities. These two compounds surpassed all the precision levels of docking and also the stringent post docking and cumulative molecular dynamics evaluations. Moreover, the backbone flexibility of hotspot residues in the hydrophobic cleft, which has been previously reported to be essential for accommodative binding of RON2 to AMA1, was also highly perturbed by these compounds. Furthermore, binding free energy calculations of these two compounds also revealed a significant affinity to AMA1. Machine learning approaches also predicted these two compounds to possess more relevant activities. Hence, these two leads, NSC95522 and NSC179676, may prove to be potential inhibitors targeting AMA1-RON2 complex formation towards combating toxoplasmosis.

• Biomolecules & Therapeutics
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• v.27 no.6
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• pp.577-583
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• 2019

Human cytochrome P450 2C9 is a highly polymorphic enzyme that is required for drug and xenobiotic metabolism. Here, we studied eleven P450 2C9 genetic variants-including three novel variants F69S, L310V, and Q324X-that were clinically identified in Korean patients. P450 2C9 variant enzymes were expressed in Escherichia coli and their bicistronic membrane fractions were prepared The CO-binding spectra were obtained for nine enzyme variants, indicating P450 holoenzymes, but not for the M02 (L90P) variant. The M11 (Q324X) variant could not be expressed due to an early nonsense mutation. LC-MS/MS analysis was performed to measure the catalytic activities of the P450 2C9 variants, using diclofenac as a substrate. Steady-state kinetic analysis revealed that the catalytic efficiency of all nine P450 2C9 variants was lower than that of the wild type P450 2C9 enzyme. The M05 (R150L) and M06 (P279T) variants showed high $k_{cat}$ values; however, their $K_m$ values were also high. As the M01 (F69S), M03 (R124Q), M04 (R125H), M08 (I359L), M09 (I359T), and M10 (A477T) variants exhibited higher $K_m$ and lower $k_{cat}$ values than that of the wild type enzyme, their catalytic efficiency decreased by approximately 50-fold compared to the wild type enzyme. Furthermore, the novel variant M07 (L310V) showed lower $k_{cat}$ and $K_m$ values than the wild type enzyme, which resulted in its decreased (80%) catalytic efficiency. The X-ray crystal structure of P450 2C9 revealed the presence of mutations in the residues surrounding the substrate-binding cavity. Functional characterization of these genetic variants can help understand the pharmacogenetic outcomes.

• Korean Journal of Veterinary Research
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• v.60 no.4
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• pp.209-213
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• 2020

This study examined the residue of dl-methylephedrine hydrochloride (MEP) on the muscle of pigs administered orally with MEP 12 g/ton feed for seven consecutive days. Twenty healthy cross swine were administered MEP. Four treated animals were selected arbitrarily to be sacrificed at 1, 2, 3, 4, and 5 days after treatment. MEP residue concentrations in the muscle were determined by liquid chromatography coupled with tandem mass spectrometry. The drug was extracted from muscle samples using 10 mM ammonium formate in acetonitrile followed by clean-up with n-hexane. The analyte was separated on an XBridgeTM hydrophilic interaction liquid chromatography column using 10 mM ammonium formate in deionized distilled water and acetonitrile. The correlation coefficient (R2) of the calibration curve was 0.9974, and the limits of detection and quantification were 0.05 and 0.15 ㎍/kg, respectively. The recoveries at three spiking levels were 94.5-101.2%, and the relative Standard Deviations was less than 4.06%. In the MEP-treated group, MEP residues on one day post-treatment were below the maximum residue limit in the muscle. The developed method is sensitive and reliable for the detection of MEP in porcine muscle tissues. Furthermore, it exhibits low quantification limits for animal-derived food products destined for human consumption.

• Journal of Microbiology and Biotechnology
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• v.32 no.4
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• pp.504-513
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• 2022

Chitin deacetylase (CDA) inhibitors were developed as novel antifungal agents because CDA participates in critical fungal physiological and metabolic processes and increases virulence in soil-borne fungal pathogens. However, few CDA inhibitors have been reported. In this study, 150 candidate CDA inhibitors were selected from the commercial Chemdiv compound library through structure-based virtual screening. The top-ranked 25 compounds were further evaluated for biological activity. The compound J075-4187 had an IC50 of 4.24 ± 0.16 µM for AnCDA. Molecular docking calculations predicted that compound J075-4187 binds to the amino acid residues, including active sites (H101, D48). Furthermore, compound J075-4187 inhibited food spoilage fungi and plant pathogenic fungi, with minimum inhibitory concentration (MIC) at 260 ㎍/ml and minimum fungicidal concentration (MFC) at 520 ㎍/ml. Therefore, compound J075-4187 is a good candidate for use in developing antifungal agents for fungi control.

• Korean Journal of Veterinary Research
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• v.62 no.2
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• pp.18.1-18.8
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• 2022

This study investigated dexamethasone (DXM) residues in the milk from intramuscularly dosed dairy cows and established the withdrawal time (WT) of DXM in milk. Eighteen healthy Holstein cows were injected with 20 (DXM-1) or 40 mL (DXM-2) of a drug containing 1 mg/mL of DXM. After administering DXM, milk samples were collected from all cows at 12-hour intervals for five days. The DXM residue concentrations in milk were determined by liquid chromatography-tandem mass spectrometry. The correlation coefficient of the calibration curve was 0.9966, and the limits of detection and quantification (LOQ) were 0.03 and 0.1 ㎍/kg, respectively. The recoveries were 97.0% to 104.0%, and the coefficient of variations was less than 7.22%. After treatment, DXM in DXM-1 was detected above the LOQ in two milk samples at 36 hours and below the LOQ in all milk samples of DXM-2 at 48 hours. Using the WT calculation program WT 1.4, the withdrawal periods of DXM-1 and DXM-2 in milk were established to be two days. In conclusion, the developed analytical method is sensitive and reliable for detecting DXM in milk. The estimated WT of DXM in bovine milk is shorter than the current milk WT recommendation of three days for DXM in lactating dairy cows.