• Bulletin of the Korean Chemical Society
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• 제28권12호
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• pp.2248-2252
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• 2007

SOMT-9 is an O-methyltransferase that utilizes quercetin to produce 3'-methoxy quercetin. In order to determine which amino acids of SOMT-9 are important for this reaction and its regioselectivity, molecular docking experiments followed by site directed mutagenesis were performed. Molecular modeling and molecular docking experiments identified several amino acid residues involved in metal binding, AdoMet binding, and substrate binding. Site-directed mutagenesis showed that Asp188 is critical for metal binding and that Lys165 assists other metal binding residues in maintaining quercetin in the proper position during the reaction. In addition, Tyr207 was shown to play an important role in the determination of the regioselectivity and Met60 was shown to be involved in formation of the hydrophobic pocket necessary for substrate binding. The molecular modeling and docking experiments discussed in this study could be applicable to future research including prediction of substrate binding and regioselectivity of an enzyme.

• Journal of Microbiology and Biotechnology
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• 제30권1호
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• pp.146-154
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• 2020

With the advantages of biocatalytic method, enzymes have been excavated for the synthesis of chiral amino acids by the reductive amination of ketones, offering a promising way of producing pharmaceutical intermediates. In this work, a robust phenylalanine dehydrogenase (PheDH) with wide substrate spectrum and high catalytic efficiency was constructed through rational design and active-site-targeted, site-specific mutagenesis by using the parent enzyme from Bacillus halodurans. Active sites with bonding substrate and amino acid residues surrounding the substrate binding pocket, 49L-50G-51G, 74M,77K, 122G-123T-124D-125M, 275N, 305L and 308V of the PheDH, were identified. Noticeably, the new mutant PheDH (E113D-N276L) showed approximately 6.06-fold increment of k_cat/Km in the oxidative deamination and more than 1.58-fold in the reductive amination compared to that of the wide type. Meanwhile, the PheDHs exhibit high capacity of accepting benzylic and aliphatic ketone substrates. The broad specificity, high catalytic efficiency and selectivity, along with excellent thermal stability, render these broad-spectrum enzymes ideal targets for further development with potential diagnostic reagent and pharmaceutical compounds applications.

• BMB Reports
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• 제43권6호
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• pp.419-426
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• 2010

Aeromonas hydrophila is a bacterial pathogen that infects a large number of eukaryotes, including humans. The UDP-galactose 4'-epimerase (GalE) catalyzes interconversion of UDP-galactose to UDP-glucose and plays a key role in lipopolysaccharide biosynthesis. This makes it an important virulence determinant, and therefore a potential drug target. Our earlier studies revealed that unlike other GalEs, GalE of A. hydrophila exists as a monomer. This uniqueness necessitated elucidation of its structure and active site. Chemical modification of the 6xHis-rGalE demonstrated the role of histidine residue in catalysis and that it did not constitute the substrate binding pocket. Loss of the 6xHis-rGalE activity and coenzyme fluorescence with thiol modifying reagents established the role of two distinct vicinal thiols in catalysis. Chemical modification studies revealed arginine to be essential for catalysis. Site-directed mutagenesis indicated Tyr149 and Lys153 to be involved in catalysis. Use of glycerol as a cosolvent enhanced the GalE thermostability significantly.

• Molecules and Cells
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• 제27권1호
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• pp.99-103
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• 2009

Ribose-5-phosphate isomerase A (RpiA) plays an important role in interconverting between ribose-5-phosphate (R5P) and ribulose-5-phosphate in the pentose phosphate pathway and the Calvin cycle. We have determined the crystal structures of the open form RpiA from Vibrio vulnificus YJ106 (VvRpiA) in complex with the R5P and the closed form with arabinose-5-phosphate (A5P) in parallel with the apo VvRpiA at $2.0{\AA}$ resolution. VvRpiA is highly similar to Escherichia coli RpiA, and the VvRpiA-R5P complex strongly resembles the E. coli RpiA-A5P complex. Interestingly, unlike the E. coli RpiA-A5P complex, the position of A5P in the VvRpiA-A5P complex reveals a different position than the R5P binding mode. VvRpiA-A5P has a sugar ring inside the binding pocket and a phosphate group outside the binding pocket: By contrast, the sugar ring of A5P interacts with the Asp4, Lys7, Ser30, Asp118, and Lys121 residues; the phosphate group of A5P interacts with two water molecules, W51 and W82.

• Genomics & Informatics
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• 제12권4호
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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.

• Bulletin of the Korean Chemical Society
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• 제29권5호
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• pp.959-962
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• 2008

We have performed FlexX docking experiments to predict the best docking poses of 5-androst-16-en-3-ol or 5-androst-16-en-3-one to Boar salivary lipocalin (SAL). Since no steroids were found inside of the binding pocket of the X-ray structure of 1GM6, we tried to find docking structures after opening the pocket using the random tweak option implemented in SYBYL. This operation allowed the ligand to enter the pocket. The best poses generated from FlexX were different from the structures reported earlier, which calculated docking poses by manual docking followed by minimization. Analysis of docking poses allowed us to identify pharmacophores. From this information, virtual screening experiments using UNITY were performed. Among six candidates, 3-(3,7-dimethyloct-6-enylamino)propane-1,2-diol (Leadquest code name: 5755) was chosen for further development. Future work will involve synthesis of some derivatives of 5755 and biological experiments if any derivatives can control the biostimulation and improve reproductive efficiency in pigs.

• 한국생물정보학회:학술대회논문집
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• 한국생물정보시스템생물학회 2005년도 BIOINFO 2005
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• pp.256-259
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• 2005

Among non-synonymous SNPs that cause amino acid change in the protein product, the selection of disease-causing SNPs has been of great interest. We present the comparison between the evolutionary (SIFT score) and structural information (binding pocket) to show that the incorporation between them provides an advantage of sorting disease-causing SNPs from normal SNPs. To set up the procedure, we apply the machine learning method to the test data set from the laboratory experiments.

• Bulletin of the Korean Chemical Society
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• 제27권1호
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• pp.59-64
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• 2006

Unlike other viral polymerases, HCV RNA-dependent RNA polymerase (RdRp) has not been successfully inhibited by nucleoside analogues presumably due to its strong substrate specificity for RNA. Thus, in order to understand the RNA-specificity of HCV RdRp, the structural characteristics of the active site was investigated. The hereto unknown 2-OH binding pocket at the active site of RdRp provides invaluable implication for the development of novel anti-HCV nucleoside analogues.

• Bulletin of the Korean Chemical Society
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• 제22권11호
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• pp.1248-1254
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• 2001

The conformations and energies of p-tert-butylcalix[4]crown-6-ether (1) and its alkyl ammonium complexes have been calculated by ab initio HF/6-31G quantum mechanics method. The cone conformation was found to be most stable for free host 1. We hav e determined the binding site of these host-guest complexes focusing on the crown-6-ether or p-tert-butylcalix[4]arene pocket of the cone conformation of host molecule 1. The primary binding site of host 1 for the recognition of alkyl ammonium guests was confirmed to be the central part of the crown moiety of cone conformation. The complexation energy calculations revealed that the ammonium cation without alkyl group showed the highest complexation efficiency when combined with host 1, that is in satisfactory agreement with the experimental results.

• BMB Reports
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• 제50권10호
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• pp.485-486
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• 2017

Many intrinsically unstructured/unfolded proteins (IUPs) contain transient local secondary structures even though they are "unstructured" in a tertiary sense. These local secondary structures are named "pre-structured motifs (PreSMos)" and in fact are the specificity determinants for IUP-target binding, i.e., the active sites in IUPs. Using high-resolution NMR we have delineated a PreSMo active site in the intrinsically unfolded mid-domain (residues 201-300) of SUMO-specific protease 4 (SUSP4). This 29-residue motif which we termed a p53 rescue motif can protect p53 from mdm2 quenching by binding to the p53-helix binding pocket in mdm2(3-109). Our work demonstrates that the PreSMo approach is quite effective in providing a structural rationale for interactions of p53-mdm2-SUSP4 and opens a novel avenue for designing mdm2-inhibiting anticancer compounds.