• Applied Biological Chemistry
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• 제33권4호
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• pp.287-292
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• 1990

황금종 대두로 부터 8종의 Bowman-Birk형 proteinase inhibitor들을 DEAE-Sephadex A-50으로 전기영동상 단일 band로 순수하게 분리하였다. 이중 inhibitor VII cysteine함량이 17%로 높고 trypsin 및 chymotrypsin에 대해 각각 독립적인 결합부위를 가지고 있으며 상기 두 효소에 대한 저해활성도의 비(TIA/CIA)가 1.0으로 전형적인 Bowman-Birk trypsin inhibitor(BBTI)로 확인되었다. 본 inhibitor와 trypsin 및 chymotrypsin complex의 dissociation constant는 각각 $9.17{\times}10^{-9}M$과 $5.14{\times}10^{-8}M$로 매우 안정하였다. 또한 inhibitor Vll은 열에 매우 안정한 단백질로 $100^{\circ}C$, 6시간 처리에도 저해활성도 감소가 50%밖에 안되었다. 순수분리된 7종의 다른 isoinhibitor중 inhibitor III만이 TIA/CIA값이 1.2로 BBTI와 비슷하였으나 그외 inhibitor I, II, IV, V,및 VIII은 그 값이 $3{\sim}29$로 BBTI와는 성질이 다른 isoinhibitnr로 추정되었다.

• 한국환경과학회지
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• 제8권3호
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• pp.337-347
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• 1999

This study was carried out to evaluate the characteristics of dissolved organics based on their origins, which were divided into two categories. The first group consisted of river, lake and secondary sewage treatment effluent, which were chosen as representative of their origins. The second group were artificial samples which were made of AHA(Aldrich humic acids) and WHA(Wako humic acids). Physicochemical characteristics, biological degradability and THMEP(trihalomethane formation potential) of the samples were analysed based on the AMWD(apparent molecular weight distribution). Large portion of dissolved organic carbon(DOC) in the river and lake samples was comprised of LMW(low molecular weight), which that of AHA and WHA was HMW(high molecular weight). The DOC of the lake was evenly distributed in the all range of molecular weight. The river, lake and secondary treated effluent have lower ultraviolet(UV) absorbance at 254nm, and have a higher amount of humic acids. Higher absorbance of humic acids means that aliphatic bond and benzenoid type components that absorb UV light were contained in these kind of humic acids. It was expected that lake sample was the most biodegradable in the different samples investigated, and in order of secondary sewage treatment effluent, river, WHA and AHA based on the result of determination of specific ultraviolet absorbance(SUVA). Biodegradability showed similar result except for AHA, while dissolved organics in the range of LMW decreased during the biodegradability test, and on the contrary those of HMW increased. Production of the SMPs(soluble micobial products) was observed during humicfication of dissolved organics and the SMPs were higher production of the SMPs. THM formation was high in the samples containing HMW and similar tendency was shown in the THMEP(trihalomethane formation potential), except for WHA.

• BMB Reports
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• 제32권3호
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• pp.239-246
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• 1999

A tripeptidase (PepT) was purified to homogeneity from Bacillus subtilis through four sequential chromatographies including DEAE-Sepharose ion exchange, hydroxylapatite, mono-Q FPLC ion exchange, and Superose-12 FPLC gel filtration. The apparent molecular mass of the enzyme was 49,200 Da and 51,400 Da as determined by sodium dodecylsulfatepolyacrylamide gel electrophoresis (SDS-PAGE) and gel filtration chromatography, respectively, and the enzyme exists in a monomeric form. The physicochemical properties of the enzyme were as follows: optimum pH at 7.5, optimum temperature at $60^{\circ}C$, and pI at 4.9. The $K_m$ and $V_{max}$ values of the enzyme were 4.3 mM and 2.5 mmol/min/mg, respectively, with MetAla-Ser as substrate. The B. subtilis PepT requires $Co^{2+}$ ion(s) for activation, while it is inactivated by EOTA and 1,10-phenanthroline, suggesting that it is a metalloprotein. The enzyme was not inhibited by any of serine protease, aspartic protease, or leucine aminopeptidase inhibitors. The enzyme showed comparable activities towards four different substrates including Met-Ala-Ser, Leu-Gly-Gly, Leu-Ser-Phe, and Leu-Leu-Tyr. The amino terminal sequence of PepT determined by Edman degradation was found to be MKEEIIERFTTYVXV and turned out to be identical to that of PepT deduced from a cloned B. subtilis pepT.

• 한국동물학회지
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• 제39권3호
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• pp.307-316
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• 1996

담배나방 (Helicoverpa assulta)의 발생기간 중 존재하는 major haemolyruph protein (MHP)을 확인하였으며, 그 발생에 따른 변화상 및 물리화학적 특성을 조사하였다. MHP는 유충-용-성충의 발생단계에서 확인되었으며, 그 함량은 유충의 성장에 따라 증가하여 용기와 성충기 동안에도 높게 유지되고 용초기와 성충 초기에서 일시적인 감소를 보여, 총 혈림프 단백질의 농도 변화와 유사한 변화양상을 보였다. ammonium sulfate precipitation, gel filtration 그리고 ion exchange chromatography를 통해 정제한 MHP의 전기영동 분석 결과, 단일 구성소 단위 (69 kDa)가 hexamer를 이룬 분자량 414kDa의 glycolipoprotein (pI 5.9)인 것으로 확인되었다. MHP의 아미노산 조성에 있어서 18.27 mole % 의 tryptophan, 7.47 mole % 의 tyrosine and 6.51 mole % 의 phenylalanine이 검출되어 다른 곤충류의 저단백질들에 비해 비교적 높은 aromatic amino acid의함량을 보였다. 지방체, 장, 말피기관 단백질의 전기영동 및 이들 단백질과 anti-MHP간의 immunodiffusion test 결과 MHP는 지방체에 존재하고 있음이 확인되었다.

• 한국자기공명학회논문지
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• 제19권2호
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• pp.74-82
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• 2015

Ryanodine receptor (RyR) is one of the two major $Ca^{2+}$ channels in membranes of intracellular $Ca^{2+}$ stores and is found in sarcoplasmic reticulum (SR), endoplasmic reticulum (ER). RyR1 is also the major calmodulin-binding protein of sarcoplasmic reticulum membranes. Residues 4064-4210 in the RyR1 polypeptide chain has similar primary sequence with calmodulin (CaM) and was designated as CaM-like domain (CaMLD). When expressed as a recombinant peptide, CaMLD showed several CaM-like properties in previous studies. Still, previous studies of CaMLD were focused on protein-protein interactions rather than its own properties. Here, we studied the expression of CaMLD and its sub-domains corresponding to each lobe of CaM in Escherichia coli. CaMLD could be obtained only as inclusion body, and it was refolded using urea solubilization followed by dialysis. Using spectroscopic approaches, such as NMR, circular dichroism, and gel filtration experiment, we found that the refolded CaMLD exists as nonspecific aggregate, even though it has alpha helical secondary structure. In comparison, the first half of CaMLD (R4061-4141) could be obtained as natively soluble protein with thioredoxin fusion. After the removal of the fusion tag, it exhibited folded and helical properties as shown by NMR and circular dichroism experiments. Its oligomeric status was different from CaMLD, existing as dimeric form in solution. However, the second half of the protein could not be obtained as soluble protein regardless of fusion tag. Based on these results, we believe that CaMLD, although similar to CaM in sequence, has quite different physicochemical properties and that the second half of the protein renders it the aggregative properties.

• Journal of Pharmaceutical Investigation
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• 제35권2호
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• pp.107-110
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• 2005

To develop a clotrimazole-loaded solid suppository with poloxamer and propylene glycol, the melting points of various formulations composed of poloxamer 188 (P 188) and propylene glycol were investigated. The dissolution study of clotrimazole delivered by the suppository composed of P 188 and propylene glycol was performed. The mixtures composed of P 188 and propylene glycol were homogeneous. Propylene glycol affected the melting points of poloxamer mixtures. In particular, the mixture [P 188/propylene glycol (70/30%)] with the melting point of about $32^{\circ}C$ was a solid form at room temperature and instantly melted at physiological temperature. Furthermore, propylene glycol affected greatly the dissolution rates of clotrimazole from the suppository. Dissolution mechanism analysis showed the dissolution of clotrimazole was proportional to the time. Our results indicated that the solid suppository with P 188 and propylene glycol would be a candidate of rectal dosage form for clotrimazole.

• Journal of Microbiology and Biotechnology
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• 제11권6호
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• pp.1011-1017
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• 2001

Streptomyces sp. Y-110 cytochrome P-450, induced by the addition of compactin -Na into the culture medium, was purified from the cell extract to apparent homogeniety, mainly by DEAE-Sepharose, hydroxyapatite, and Mono Q column chromatyography. The sepcific activity of purified enzyme on its substrate, compactin-Na, was determined to be 15 nmol of pravastatin per mg protein. The molecular mass of this enzyme on SDS-PAGE was $37{\pm}0.5$ kDa, pI was 4.5, and its CO difference spectrum showed maximum absorption peaks at 452 and 550nm, respectively. The N-terminal amino acid sequence was determined to be Met>Thr>Cys>Thr>Pro>Val>Thr>Val>The>Gly>Ala>Ala>Gly>Gln>Ile>Gly>Tyr>Ala>Leu. Its apparent $K_m$ on compactin-Na was $1.294{\mu}M{\cdot}min^-1,\;and\;V_{max}\;was\;1.028{\mu}M{\cdot}min^-1$. The maximum substrate concentration ($K_s$) for reaction was $270 {\mu}M$and thus $1/[K_s]$ was $3.7{\mu}M$. These physicochemical characteristics and kinetic behavior of this enzyme were compared and shown to be different from those of Streptomyces cytochrome P-450 enzymes reported, suggesting that this enzyme may be an additional member of the Streptomyces cytochrome P-450 family.

• Asian Pacific Journal of Cancer Prevention
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• 제15권23호
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• pp.10281-10287
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• 2015

Background: Development of a nanosized polymeric delivery system for erlotinib was the main objective of this research. Materials and Methods: Poly caprolactone-polyethylene glycol-polycaprolactone (PCEC) copolymers with different compositions were synthesized via ring opening polymerization. Formation of triblock copolymers was confirmed by HNMR as well as FT-IR. Erlotinib loaded nanoparticles were prepared by means of synthesized copolymers with solvent displacement method. Results: Physicochemical properties of obtained polymeric nanoparticles were dependent on composition of used copolymers. Size of particles was decreased with decreasing the PCL/PEG molar ratio in used copolymers. Encapsulation efficiency of prepared formulations was declined by decreasing their particle size. Drug release behavior from the prepared nanoparticles exhibited a sustained pattern without a burst release. From the release profiles, it can be found that erlotinib release rate from polymeric nanoparticles is decreased by increase of CL/PEG molar ratio of prepared block copolymers. Based on MTT assay results, cell growth inhibition of erlotinib has a dose and time dependent pattern. After 72 hours of exposure, the 50% inhibitory concentration (IC50) of erlotinib hydrochloride was appeared to be $14.8{\mu}M$. Conclusions: From the obtained results, it can be concluded that the prepared PCEC nanoparticles in this study might have the potential to be considered as delivery system for erlotinib.

• Journal of Pharmaceutical Investigation
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• 제41권2호
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• pp.95-102
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• 2011

An amphiphilic cationic branched methoxy poly (ethylene glycol)-branched polyethylenimine - poly(L-phenylalanine) (mPEG-bPEI-pPhe) block copolymer was successfully synthesized by ring-opening polymerization (ROP) of N-carboxyanhydride of L-phenylalanine (Phe-NCA) with mPEG-bPEI for the preparation of more stable polyelectrolyte complex (PEC) included a hydrophobic interaction. mPEG-bPEI was firstly prepared by the coupling of mPEG and bPEI using hexamethylene diisocyanate (HMDI). The structural properties of mPEG-bPEI-pPhe copolymers were confirmed by $^1H$ NMR. The copolymers exhibited a self-assemble behavior in water above critical aggregate concentration (CAC) in the range of 0.01-0.14 g/L. The CAC of copolymers obviously depended on the hydrophobic block content in the copolymers (the value decreased with the increase of the pPhe block content). The cationic copolymers have the ability to form multi-interaction complex (MIC) with bovine serum albumin (BSA) and plasmid DNA through multi-interaction (electrostatic and hydrophobic interaction). The physicochemical characterization of the complex was carried out by the measurement of zeta potential and particle size. Their zeta-potentials were positive (approximately +10 mV) and their sizes decreased with increasing pPhe contents in the copolymers (PPF/BSA wt% ratio = 2). The complex showed good stability at high ionic strength. Therefore, mPEG-bPEI-pPhe block copolymer was considered as a potential material to enhance the stability of complex including biotherapuetic drugs.

• Journal of Microbiology and Biotechnology
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• 제18권2호
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• pp.248-254
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• 2008

Candida magnoliae, an osmotolerant and erythritol producing yeast, prefers D-fructose to D-glucose as carbon sources. For the investigation of the fructophilic characteristics with respect to sugar transportation, a sequential extraction method using various detergents and ultracentrifugation was developed to isolate cellular membrane proteins in C. magnoliae. Immunoblot analysis with the Pma1 antibody and two-dimensional electrophoresis analysis coupled with MS showed that the fraction II was enriched with membrane proteins. Eighteen proteins out of 36 spots were identified as membrane or membrane-associated proteins involved in sugar uptake, stress response, carbon metabolism, and so on. Among them, three proteins were significantly upregulated under the fructose supplying conditions. The hexose transporter was highly homologous to Ght6p in Schizosaccharomyces pombe, which was known as a predominant transporter for the fructose uptake of S. pombe because it exhibited higher affinity to D-fructose than D-glucose. The physicochemical properties of the ATP-binding cassette transporter and inorganic transporter explained their direct or indirect associations with the fructophilic behavior of C. magnoliae. The identification and characterization of membrane proteins involved in sugar uptake might contribute to the elucidation of the selective utilization of fructose to glucose by C. magnoliae at a molecular level.