• Toxicological Research
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• v.23 no.1
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• pp.1-9
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• 2007

Transporters are membrane proteins that mediate the transfer of substrate across the cellular membrane. In this overview, the characteristics and the toxicological relevance were discussed for various types of transporters. For drug transporters, the overview focused on ATP-binding cassette transporters and solute carrier family 21A/22A member transporters. Except for OCTN transporters and OATP transporters, drug transporters tend to have broad substrate specificity, suggesting drug-drug interaction at the level of transport processes (e.g., interaction between methotrexate and non-steroidal anti-inflammatory agents) is likely. For metal transporters, transporters for zinc, copper and multiple metals were discussed in this overview. These metal transporters have comparatively narrow substrate specificity, except for multiple metal transporters, suggesting that inter-substrate interaction at the level of transport is less likely. In contrast, the expressions of the transporters are often regulated by their substrates, suggesting cellular adaptation mechanism exists for these transporters. The drug-drug interactions in drug transporters and the cellular adaptation mechanisms for metal transporters are likely to lead to alterations in pharmacokinetics and cellular metal homeostasis, which may be linked to the development of toxicity. Therefore, the transporter-mediated alterations may have toxicological relevance.

• BMB Reports
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• v.45 no.11
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• pp.665-670
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• 2012

The insect midgut epithelium is generally lined with a unique chitin and protein structure, the peritrophic membrane (PM), which facilitates food digestion and protects the gut epithelium. We used gel electrophoresis and mass spectrometry to identify the extracted proteins from the silkworm PM to obtain an in-depth understanding of the biological function of the silkworm PM components. A total of 305 proteins, with molecular weights ranging from 8.02 kDa to 788.52 kDa and the isoelectric points ranging from 3.39 to 12.91, were successfully identified. We also found several major classes of PM proteins, i.e. PM chitin-binding protein, invertebrate intestinal mucin, and chitin deacetylase. The protein profile provides a basis for further study of the physiological events in the PM of Bombyx mori.

• Journal of Microbiology and Biotechnology
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• v.19 no.12
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• pp.1612-1616
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• 2009

Isoflavonoids are a class of phytoestrogens. Isoflavonone synthase (IFS) is responsible for the conversion of naringenin to genistein. IFS is a cytochrome P450 (CYP), and requires cytochrome P450 reductase (CPR) for its activity. Additionally, the majority of cytochrome P450s harbor a membrane binding domain, making them difficult to express in Escherichia coli. In order to resolve these issues, we constructed an inframe fusion of the IFS from red clover (RCIFS) and CPR from rice (RCPR) after removing the membrane binding domain from RCIFS and RCPR. The resultant fusion gene, RCIFS-RCPR, was expressed in E. coli. The conversion of naringenin into genistein was confirmed using this E. coli transformant. Following the optimization of the medium and cell density for biotransformation, $60\;{\mu}M$ of genistein could be generated from $80\;{\mu}M$ of naringenin. This fusion protein approach may be applicable to the expression of other P450s in E. coli.

• Bulletin of the Korean Chemical Society
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• v.32 no.9
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• pp.3219-3222
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• 2011

Three types of ${\beta}$-ketoacyl acyl carrier protein synthase (KAS) are important for overcoming the bacterial resistance problem. Recently, we reported the discovery of a antimicrobial flavonoid, YKAF01 (3,6-dihydroxyflavone), which exhibits antibacterial activity against Gram-positive bacteria through inhibition of ${\beta}$-ketoacyl acyl carrier protein synthase III (KAS III). In this report, we suggested that YKAF01 can be an inhibitor ${\beta}$-ketoacyl acyl carrier protein synthase I (KAS I) with dual inhibitory activity for KAS I as well as KAS III. KAS I is related to the elongation of unsaturated fatty acids in bacterial fatty acid synthesis and can be a good therapeutic target of designing novel antibiotics. We performed docking study of Escherichia coli KAS I (ecKAS I) and YKAF01, and determined their binding model. YKAF01 binds to KAS I with high binding affinity ($2.12{\times}10^6$) and exhibited an antimicrobial activity against the multidrug-resistant E. coli with minimal inhibitory concentration (MIC) value of 512 ${\mu}g$/mL. Further optimization of this compound will be carried out to improve its antimicrobial activity and membrane permeability against bacterial cell membrane.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.49 no.2
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• pp.137-145
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• 2016

This study screened the biological activity of an acidified gill extract of the Manila clam Ruditapes philippinarum including antimicrobial, hemolytic, membrane permeabilization, and DNA-binding activity, and purified the antimicrobial material. The acidified gill extract showed potent antimicrobial activity against Bacillus subtilis and Escherichia coli without significant hemolytic activity, but showed no membrane permeabilization or DNA-binding ability. An antimicrobial material was purified from the acidified gill extract using C₁₈ reversed-phase and cation-exchange high-performance liquid chromatography (HPLC). Treatment of the purified material with trypsin completely abolished all of the antibacterial activity against Bacillus subtilis, suggesting that the purified material is a proteinaceous antibiotic. The molecular weight of the purified material was 2571.9 Da, but no primary structural information was obtained due to N-terminal blocking. A future study should confirm the primary structure. Our results suggest that the Manila clam gill contains proteinaceous antibiotics that have a role in first-line defense. This information could be used to better understand the Manila clam innate immune system.

• Archives of Pharmacal Research
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• v.23 no.6
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• pp.535-547
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• 2000

Recent findings indicate that mechanical strain (deformation) exerted by the extracellular matrix modulates activation of airway smooth muscle cells. Furthermore, cytoskeletal organization in airway smooth muscle appears to be dynamic, and subject to modulation by receptor activation and mechanical strain. Mechanosensitive modulation of crossbridge activation and cytoskeletal organization may represent intracellular feedback mechanisms that limit the shortening of airway smooth muscle during bronchoconstriction. Recent findings suggest that receptor-mediated signal transduction is the primary target of mechanosensitive modulation. Mechanical strain appears to regulate the number of functional G-proteins and/or phospholipase C enzymes in the cell membrane possibly by membrane trafficking and/or protein translocation. Dense plaques, membrane structures analogous to focal adhesions, appear to be the primary target of cytoskeletal regulation. Mechanical strain and receptor-binding appear to regulate the assembly and phosphorylation of dense plaque proteins in airway smooth muscle cells. Understanding these mechanisms may reveal new pharmacological targets for control1ing airway resistance in airway diseases.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1996.04a
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• pp.173-173
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• 1996

포도당 수송을 증가시키는 것으로 밝혀진 brazilin의 작용이 지방세포의 단백질과 결합하여 나타나는지 알아보기 위한 실험으로 brazilin의 단백결합 성질을 살펴보았다. Brazilin의 단백결합 실험에는 [$^3$H]-brazilin을 사용하여 결합된 양을 방사선 동위원소량으로 확인하였다. 먼저 일반적인 단백결합을 알아보기 위하여 산침전을 이용한 brazilin의 BSA에 대한 결합을 살펴보았다. 그 결과 brazilin은 시간의 경과에 따라 BSA에 결합하는 양이 증가하여 2시간에 최고치에 달했으며 그 양은 약 80% 정도였다. 세포내 단백과의 결합은 BSA에 대한 결합과는 달리 1시간만에 최고치에 달하였다. 세포내 분획에 분포하는 brazilin의 양을 알기 위하여 brazilin처리 후 세포내 분획을 원심분리로 얻어 brazilin의 양을 확인하였다. 그 결과 brazilin은 cytosol에 대부분 존재하며 microsomal membrane과 nuclear에 많은 양이 존재하는 것으로 확인되었다. SDS-PAGE를 이용하여 brazilin과 결합한 각 분획의 단백질을 살펴본 결과 brazilin은 cytosol의 30, 65KD, microsomal membrane의 45, 130KD, nucear의 26, 35KD, plasma membrane의 46, 228KD protein과 반응하는 것으로 확인되었다.

• Journal of Environmental Science International
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• v.7 no.1
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• pp.62-66
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• 1998

Toxic Mastoparan B(MP-B) which is purified from the venom of the hornet Vespa basalis is a cationic amphlphilic tetradecapeptide. MP-B and Its Ala-substituted analogues were synthesized by solld phase method and the toxic peptide-membrane interactions were examined by circular dichroism(CD) spectra, fluorescence spectra, and leakage abilities in phospholipid membranes. In the presence of phospholipid vesicles, synthetic MP-B and its analogues formed amphiphilic -helical structures, but in the buffer soletion, those exhibited random coil conformation as measured by CD. Fluorescence spectra of MP-B and its analogues which indicated the binding affinity of peptide on phospholipid vesicles showed that the replacement of Lys at position 2 and 11 with Ala caused a remarkable effect in the blue shalt and that at position 2, in the leakage ability of the peptide.

• Proceedings of the Korean Biophysical Society Conference
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• 1996.07a
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• pp.37-37
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• 1996

Tenecin fragments are antimicrobial and antifungal peptide from Tenebrio molitor with highly positive charged amino acid residues. To elucidate their membrane selectivity and molecular mechanism, various forms of tenecin fragments were synthesized, and their interaction with acidic phospholipid, Gram (+), fungal and human erythrocyte membrane were investigated by ANTS/DPX leakage, membrane binding and fusion assay. (omitted)

• BMB Reports
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• v.40 no.5
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• pp.783-790
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• 2007

Receptor binding plays an important role in determining host specificity of the Bacillus thuringiensis Cry $\delta$-endotoxins. Mutations in domains II and III have suggested the participation of certain residues in receptor recognition and insect specificity. In the present study, we expressed the cloned domain II-III fragment of Cry4Ba and examined its binding characteristics to mosquito-larval midgut proteins. The 43-kDa Cry4Ba-domain II-III protein over-expressed in Escherichia coli as inclusion bodies was only soluble when carbonate buffer, pH 10.0 was supplemented with 4M urea. After renaturation via stepwise dialysis and subsequent purification, the refolded domain II-III protein, which specifically reacts with anti Cry4Ba-domain III monoclonal antibody, predominantly exists as a $\beta$-sheet structure determined by circular dichroism spectroscopy. In vitro binding analysis to both histological midgut tissue sections and brush border membrane proteins prepared from susceptible Aedes aegypti mosquito-larvae revealed that the isolated Cry4Ba-domain II-III protein showed binding functionality comparable to the 65-kDa full-length active toxin. Altogether, the data present the 43-kDa Cry4Ba fragment comprising domains II and III that was produced in isolation was able to retain its receptor-binding characteristics to the target larval midgut proteins.