• KSBB Journal
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• 제22권6호
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• pp.371-377
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• 2007

Polyhydroxyalkanoates (PHAs) are a family of microbial polyesters that can be produced by fermentation from renewable resources. PHAs can be used as completely biodegradable plastics or elastomers. In this paper, novel applications of PHAs in biosensor are described. A general platform technology was developed by using the substrate binding domain (SBD) of PHA depolymerase as a fusion partner to immobilize proteins of interest on PHA surface. It could be shown that the proteins fused to the SBD of PHA depolymerase could be specifically immobilized onto PHA film, PHA microbead, and microcontact printed PHA surface. We review the results obtained for monitoring the specific interaction between the SBO and PHA by using enhanced green fluorescent protein, red fluorescent protein, single chain antibody against hepatitis B virus preS2 surface protein and severe acute respiratory syndrome coronavirus surface antigen as model proteins. Thus, this system can be efficiently used for studying protein-protein and possibly protein-biomolecule interactions for various biotechnological applications.

• Reproductive and Developmental Biology
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• 제30권3호
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• pp.169-174
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• 2006

To search of a new porcine pheromonal odorants, the comparative molecular similarity indices analysis(CoMSIA) between porcine odorant binding protein(pOBP) as receptor and ligands of green odorants 2-(Cyclohexyloxy)tetrahydrofurane derivatives as substrate molecule were conducted and disscused quantitatively. In the optimized CoMSIA model(I-AI) with chirality($I:\;C_{1'}(R),\;C_2(S)$) in substrate molecules and atom based fit alignment(AE) of the odorants the statistical PLS results showed the best predictability of the binding affinities based on the LOO cross-validated value ${r^2}_{cv.}\;(q^2=0.856)$ and non cross-validated conventional coefficient(${r^2}_{ncv.}=0.964)$). The structural distinctions of the highest active molecules were able to understand from the interaction between pOBP and green odorants in the contour maps with CoMSIA model.

• Proceedings of the Korean Vacuum Society Conference
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• 한국진공학회 2013년도 제45회 하계 정기학술대회 초록집
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• pp.249-249
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• 2013

Surface-enhanced Raman spectroscopy (SERS) is a sensitive approach to detect and to identify a variety of molecules. To enhance the Raman signal, optimization of the gap between nanostructures is quite important. One-dimensional materials such as nanowires, nanotubes, and nanograsses have great potential to be used in SERS due to their unique sizes and shape dependent characteristics. In this study we investigate a simple way to fabricate SERS substrates based on randomly grown copper oxide (CuO) nanowires. CuO nanograss is fabricated on pre-cleaned Cu foils. Cu oxidized in an ammonium ambient solution of 2.5 M NaOH and 0.1 M $(NH_4)_2S_2O_8$ at $4^{\circ}C$ for 10, 30, and 60 minutes. Then, Cu(OH)2 nanostructures are formed and dried at $180^{\circ}C$ for 2 h. With the drying process, the Cu(OH)2 nanostructure is transformed to CuO nanograss by dehydration reaction. CuO nanograss are grown randomly on Cu foil with the average length of 10 ${\mu}m$ and the average diameter of a 100 nm. CuO nanograsses are covered by Ag with various thicknesses from 10 to 30 nm using a thermal evaporator. Then, we immerse uncoated and Ag coated CuO nanowire samples of various oxidation times in a 0.001M methanol-based 4-mercaptopyridine (4-Mpy) in order to evaluate SERS enhancement. Raman shift and SERS enhancement are measured using a Raman spectrometer (Horiba, LabRAM ARAMIS Spectrometer) with the laser wavelength of 532 nm. Raman scattering is believed to be enhanced by the interaction between CuO nanograss and Ag island film. The gaps between Ag covered CuO nanograsses are diverse from <10 nm at the bottom to ~200 nm at the top of nanograsses. SERS signal are improved where the gaps are minimized to near 10s of nanometers. There are many spots that provide sufficiently narrow gap between the structures on randomly grown CuO nanograss surface. Then we may find optimal enhancement of Raman signal using the mapping data of average results. Fabrication of CuO nanograss based on a solution method is relatively simple and fast so this result can potentially provide a path toward cost effective fabrication of SERS substrate for sensing applications.

• Animal Bioscience
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• 제34권5호
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• pp.867-879
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• 2021

Objective: Fibronectin 3 (FN3) and immunoglobulin like modules (Ig) are usually collocated beside modular cellulase catalytic domains. However, very few researches have investigated the role of these modules. In a previous study, we have sequenced and analyzed bacterial metagenomic DNA in Vietnamese goats' rumen and found that cellulase-producing bacteria and cellulase families were dominant. In this study, the properties of modular cellulases and the role of a FN3 in unique endoglucanase belonging to glycosyl hydorlase (GH) family 5 were determined. Methods: Based on Pfam analysis, the cellulases sequences containing FN3, Ig modules were extracted from 297 complete open reading frames (ORFs). The alkaline, thermostability, tertiary structure of deduced enzymes were predicted by AcalPred, TBI software, Phyre2 and Swiss models. Then, whole and truncated forms of a selected gene were expressed in Escherichia coli and purified by His-tag affinity column for assessment of FN3 ability to enhance enzyme activity, solubility and conformation. Results: From 297 complete ORFs coding for cellulases, 148 sequences containing FN3, Ig were identified. Mostly FN3 appeared in 90.9% beta-glucosidases belonging to glycosyl hydrolase family 3 (GH3) and situated downstream of catalytic domains. The Ig was found upstream of 100% endoglucanase GH9. Rarely FN3 was seen to be situated downstream of X domain and upstream of catalytic domain endoglucanase GH5. Whole enzyme (called XFN3GH5 based on modular structure) and truncate forms FN3, XFN3, FN3GH5, GH5 were cloned in pET22b (+) and pET22SUMO to be expressed in single and fusion forms with a small ubiquitin-related modifier partner (S). The FN3, SFN3 increased GH5 solubility in FN3GH5, SFN3GH5. The SFN3 partly served for GH5 conformation in SFN3GH5, increased modules interaction and enzyme-soluble substrate affinity to enhance SXFN3GH5, SFN3GH5 activities in mixtures. Both SFN3 and SXFN3 did not anchor enzyme on filter paper but exfoliate and separate cellulose chains on filter paper for enzyme hydrolysis. Conclusion: Based on these findings, the presence of FN3 module in certain cellulases was confirmed and it assisted for enzyme conformation and activity in both soluble and insoluble substrate.

• Biomolecules & Therapeutics
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• 제17권3호
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• pp.299-304
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• 2009

Rosiglitazone maleate (RGM) is widely used for improving insulin resistance. RGM is a moderate inhibitor of cytochrome P450 2C8 (CYP2C8) and is also mainly metabolized by CYP2C8. The aim of this study was to determine whether the effect of RGM on CYP2C8 is altered by co-treatment with other drugs, and whether amlodipine camsylate (AC) changes the pharmacokinetics (PK) of RGM. Of the 11 drugs that are likely to be co-administered with RGM in diabetic patients, seven drugs lowered the $IC_{50}$ value of RGM on CYP2C8 by more than 80%. In vitro CYP2C8 inhibitory assays of RGM in combination with drugs of interest showed that the $IC_{50}$ of RGM was decreased by 98.9% by AC. In a pharmacokinetic study, Sprague-Dawley (SD) rats were orally administered 1 mg/kg of RGM following by single or 10-consecutive daily administrations of 1.5 mg/kg/day of AC. No significant changes in the pharmacokinetic parameters of RGM were observed after a single administration of AC, but the AUC and $C_{max}$ values of RGM were significantly reduced by 36% and 31%, respectively, by multiple administrations of AC. In conclusion, RGM was found to be affected by AC by in vitro CYP2C8 inhibition testing, and multiple dosing of AC appreciably changed the pharmacokinetics of RGM. These findings suggest that a drug interaction exists between AC and RGM.

• Journal of Korean Society of Environmental Engineers
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• 제22권10호
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• pp.1869-1879
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• 2000

A series of experiments were conducted for modeling the fate and effect of the coupled oxidation reduction reaction of ethanol and propionate recognized as important intermediates in anaerobic degradation metabolism. Anaerobic kinetics for conversion of propionate and the interaction with ethanol were investigated using the model of specific substrate priority utilization effect. Seed cultures for the experiment were obtained from an anaerobically enriched steady-state propionate master culture reactor (HPr-MCR), ethanol-propionate master culture reactor (EtPr-MCR) and glucose master culture reactor (Glu-MCR). Experiments were consisted of four phases. Phase I, II and III were conducted by fixing the propionate organic loading as 1.0 g COD/L with increasing ethanol loading of 0, 100, 200, 400 and 1,000 mg/L, to find metabolic interaction of ethanol and propionate degradation by each enriched anaerobic culture. In phase IV, different mixing ratios of Glu-MCR and HPr-MCR cultures with fixed propionate organic loading, 1.0 g COD/L, were applied to observe the propionate degradation metabolic behavior. In the results of this study, different pathways of propionate and ethanol conversion were found using a modified competitive inhibition kinetic model. Increase of $K_{s2}$ value reflected the formation of acetate followed by ethanol degradation. In addition. $K_3$ value was increased slightly as the reactions of acetate formation and degradation were occurred in acetoclastic methanogenesis.

• Korean Journal of Microbiology
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• 제48권2호
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• pp.79-86
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• 2012

ErmSF is one of the Erm family proteins which catalyze S-adenosyl-$_L$-methionine dependent modification of a specific adenine residue (A2058, E. coli numbering) in bacterial 23S rRNA, thereby conferring resistance to clinically important macrolide, lincosamide and streptogramin B ($MLS_B$) antibiotics. $^{222}FXPXPXVXS^{230}$ (ErmSF numbering) sequence appears to be a consensus sequence among the Erm family. This sequence was supposed to be involved in direct interaction with the target adenine from the structural studies of Erm protein ErmC'. But in DNA methyltarnsferase M. Taq I, this interaction have been identified biochemically and from the complex structure with substrate. Arginine 223 and 227 in this sequence are not conserved among Erm proteins, but because of the basic nature of residues, it was expected to interact with RNA substrates. Two amino acid residues were replaced with Ala by site-directed mutagenesis. Two mutant proteins still maintained its activity in vivo and resistant to the antibiotic erythromycin. Compared to the wild-type ErmSF, R223A and R227A proteins retained about 50% and 88% of activity in vitro, respectively. Even though those arginine residues are not essential in the catalytic step, with their positive charge they may play an important role for RNA binding.

• Korean Journal of Food Science and Technology
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• 제35권5호
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• pp.791-796
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• 2003

Polyphenol oxidase isoforms were purified from the lotus roots using 50% acetone precipitation, conventional chromatographies of Q-Sepharose and hydrophobic interaction, and high performance liquid chromatographies of Mono-Q and Superdex 75 gel-filtration. Molecular mass of a purified PPO isoform (LPIII-2) was determined to be 56 kDa using gel-filtration chromatography. The active form of LPIII-2 appeared to bea heterodimer, as purified LPIII-2 on SDS-PAGE gel showed two bands that were determined to be 28 kDa and 26 kDa. To further characterize PPO, partially purified PPO isoforms (LP-II, LP-III) were obtained from Q-Sepharose anion-exchange chromatography. In substrate specificity, the partially purified PPO isoform LP-II showed a high affinity to catechol, while LP-III showed a high affinity to pyrogallol. The optimum pH of LP-II and LP-III was pH 7.0. Interestingly, the partially purified PPO isoforms showed high activities at low temperatures $(0{\sim}5^{\circ}C)$, and as temperatures rose, the activities decreased. Both PPO isoforms were stable at $40^{\circ}C$ and were inactivated by incubation at $60^{\circ}C$ for 40 min.

• KSBB Journal
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• 제24권3호
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• pp.227-238
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• 2009

In the recent years, some organic toxic chemicals were used for obtaining high-yield productivity in agriculture. The undegraded pesticides may remain in the agricultural foods through atmosphere, water, and soil and cause public health problems to environmental resources and human beings even at very low concentrations. Small amounts of pesticides can affect a central nervous system, resulting in immunogenic diseases, infertility problems, respiratory diseases and born marrow diseases, which can lead even to death. Monitoring of the environmental pesticide is one of the important issues for the human well-being. Several kinds of biosensors have been successfully applied to the detection of agrichemical toxicity. Also, few platforms for biocide detection have been definitely developed for the degradation and reaction of pesticides. Biochip and electrochemistry experiments involve immobilizing a receptor molecule on a solid substrate surface, and monitoring its interaction with an analyze in a sample solution. Furthermore, nanotechnology can be applied to make high-throughput analyses that are smaller, faster and sensitive than conventional assays. Some nanomaterials or nanofabricated surfaces can be coupled to biomolecules and used in antibody-based assays and enzymatic methods for pesticide residues. The operation procedure has become more convenient as it does not require labeling procedure. In this paper, we review the recent advances in agrichemical defection research and also describe the label-free biosensor for pesticides using various useful detection methods.

• Journal of the Korean Chemical Society
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• 제54권5호
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• pp.567-572
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• 2010

5,10-Methenyltetrahydrofolate synthetase from chicken liver was purified through 30-70% ammonium sulfate fractionation, Q Sepharose Fast Flow anion exchange and Source 15Phe hydrophobic interaction chromatography. Specific activities of cell extract, ammonium sulfate, Q Sepharose Fast Flow and Source 15Phe were 0.0085, 0.031, 0.80 and 1.27 U/mg, respectively. Purification fold activities of cell extract, ammonium sulfate, Q Sepharose Fast Flow and Source 15Phe were 1, 3.7, 94.1 and 149.4, respectively. HPLC gel permeation chromatography and SDS-polyacrylamide electrophoresis experiments indicated that the enzyme is a monomeric protein with a molecular weight of 22.8 kDa. Km for 5-methyl THF and Mg-ATP were $7.1\;{\mu}M$ and $63\;{\mu}M$, respectively. Optimum temperature and pH were $30^{\circ}C$ and 6.0, respectively. The data for metal ion specificity and stoichiometry showed that the maximum activity was obtained with a 1:l. ratio of $Mg^{2+}$. The ATP and Km values increased in the order of MgATP, MgCTP, MgUTP and MgGTP, and the maximum activities also decreased in the same order, indicating MgATP as the most efficient substrate. The enzyme was chemically modified only by tetranitrometane and 1-ethyl-3-(3-dimethyl aminopropyl)-carbodiimide, indicating that tyrosine and carboxylate are present in the active site.