• 대한약학회:학술대회논문집
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• 대한약학회 2000년도 NEW STRATEGY FOR DRUG DEVELOPMENT IN POST-GENOMIC ERA(대한약학회)
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• pp.269.2-270
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• 2000

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1994년도 춘계학술대회 and 제3회 신약개발 연구발표회
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• pp.264-264
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• 1994

본 연구는 보리 씨앗에 존재하는 리보솜 불활성화 단백질(RIP) 의 삼차원 구조를 X-선 결정학 방법을 이용하여 밝히고, 그 결과로 분자 차원에서 기능을 이해하는 것을 목적으로 하고있다. 리보솜불활성화 단백질은 N-glucosidase 반응을 통하여 단백질 합성을 저해하기 때문에 세포를 죽일 수 있다. 따라서 암세포만을 특정적으로 인식하는 다른 물질과 결합시키면 암세포만을 특정적으로 죽일 수 있는 면역독소로 이용될 수 있다. 또, 최근에는 항바이러스의 작용을 함이 밝혀져 많은 연구가 진행되고 있다. 단백질 삼차원 구조 규명을 위해서는 여러가지 단계가 있는데 지난 번 과제까지 성공적으로 리보솜 불활성화 단백질의 대량 분리와 X-선 결정학의 필수 요건 좋은 결정을 길렀고, 이번에는 구조 해석을 위해 꼭 해결해야하는 위상문제를 극복하기 위하여 여러가지 실험을 진행하였다. 우선, 비슷한 구조인 피마자씨에서 분리한 Ricin의 A-체인과 미국자리공 잎에서 분리한 Pokeweed antiviral protein의 삼차원 분자좌표를 이용하여 분자치환법으로 위상문제 해결을 시도하였다. Ricin 의 A-체인을 이용하였을 때 분자의 위치가 정확히 찾아지지 않았고, 다른 모델인 Pokeweed antiviral protein을 이용하여 X-PLOR 프로그램내의 PC refinement법으로 분자치환을 시도하였다. Euler각도로 (187.37, 22.5, 311.94) 의 회전해 (Rotation solution) 를 가지고 있었고, 이러한 해에 맞추어서 분자를 돌려둔 후, 이동해 (Transaltion solution) 을 구해서 그 위치 (Orientation) 로 분자를 이동하였다. 이 때 R값은 53.9 % (8.0 - 3.5$\AA$) 이였고, 부분좌표 (Fractional coorcdinate) 에서는 0.102, 0.000, 0.261 이고, 직교좌표 (Orthogonal coorclinate) 에서는 4.616, 0.000, 13.167 의 결과를 얻었다.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1996년도 춘계학술대회
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• pp.277-277
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• 1996

Anti-CALLA Fab-RTA immunotoxins were constructed using two crosslinking agent, SMPT and SMCC, to generate a disulfide and a thioether bridge between Fab fragment of K269-65 MoAb and RTA toxin moieties, respectively. These immunotoxins were selectively immunoreative with CALLA$\^$+/ B-lineage Daudi cells. SMPT and SMCC mediated RTA immunotoxins were prepared with 49% and 53% of the RTA conjugation yields, respectively. SDS-PAGE analysis show that immunotoxins were constructed with major Fab-1 RTA (76kda), minor Fab-2RTA (106kda) and Fab-3RTA (136kda) compositions. The breakdown rates of immunotoxins were determined in the presence of glutathione by measuring the amount of reduced immunotoxins using size-exclusion HPLC. The SMCC immunotoxins were more resistant to the glutathione than SMPT immunotoxins. But, our data showed that the SMPT mediated disulfide bonded immunotoxins were much more active than the SMCC mediated thioether bonded immunotoxins to kill the target cells in vitro.

• Bulletin of the Korean Chemical Society
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• 제16권12호
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• pp.1153-1162
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• 1995

The conformational details of β-lactose are investigated through molecular dynamics simulations in conjunction with the adiabatic potential energy map. The adiabatic energy map generated in vacuo contains five local minima. The lowest energy structure on the map does not correspond to the structure determined experimentally by NMR and the X-ray crystallography. When aqueous solvent effect is incorporated into the energy map calculation by increasing the dielectric constant, one of the local minima in the vacuum energy map becomes the global minimum in the resultant energy map. The lowest energy structure of the energy map generated in aquo is consistent with the one experimentally determined. Molecular dynamics simulations starting from those fivelocal minima on the vacuum energy map reveal that conformational transitions can take place among various conformations. Molecular dynamics simulations of the lactose and ricin B chain complex system in a stochastic boundary indicate that the most stable conformation in solution phase is bound to the binding site and that there are conformational changes in the exocyclic region of the lactose molecule upon binding.

• Asian-Australasian Journal of Animal Sciences
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• 제23권10호
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• pp.1308-1318
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• 2010

The purpose of this study was to evaluate the effect of replacing soybean meal with treated castor meal with (CMT) or without lime (CMNT) on the nutrient intake, performance, carcass characteristics, and yield of commercial cuts of beef cattle from a feedlot. Thirty male, castrated, crossbreed zebu cattle were used in the study, with an average initial weight of $360{\pm}30.27\;kg$. Five animals were used as a control group and were slaughtered at the beginning of the experiment; the remaining animals (n = 25) were distributed in random blocks (repetitions), with body weight as the criterion for block assignment. The animals were fed a diet containing 65% corn silage and 35% of concentrate on dry matter (DM) basis. Five diets consisted of four levels of soybean meal (SM) substituted with CMT (0, 33, 67 and 100%) on a DM basis and a diet with 100% of SM replaced with CMNT. At the end of the experiment, all animals were slaughtered, and their gastrointestinal tracts were emptied to determine their empty body weights (EBW). No significant effects were observed (p>0.05) for the substitution of soybean meal with CMT on intake of dietary nutrients, the average daily body weight gain (ADG) or EBW gain (EBWG). In spite of greater (p<0.05) ricin intake for the diet containing CMNT (3.06 mg/kg BW) compared to the CMT diet (0.10 mg/kg BW/d), there were no effects (p>0.05) on intake of dietary nutrients, ADG or EBWG. The average intake of DM and the ADG were 10,664.63 and 1,353.04 g/d, respectively. Regarding carcass characteristics, only carcass yield in relation to body weight was linearly reduced (p<0.05) upon substitution of SM by CMT. There was no effect (p>0.05) of the substitution of SM by CMT or CMNT on the yield of carcass basic cuts. CMT prices that are higher than 85% of the SM price do not economically justify the use of CMT. For CMT prices between 20 and 80% of the SM price, the optimal level was 67% substitution, while for prices below 15% of the SM price, the optimal level was 100% substitution with CMT. It can be concluded that treated castor meal with 6% lime can totally replace soybean meal in beef cattle diets.

• 대한수의학회지
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• 제37권3호
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• pp.541-546
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• 1997

VT2(Verotoxin-2)의 효소활성 영역에 해당되는 두 영역의 아미노산들에 대하여, 첫번째 보존영역의 Glu167을 conservative point mutation 시키고, 두 번째 보존영역의 구성 아미노산 5개 전부를 deletion mutation 시켜, 각 변이주에서 독성의 감소 정도를 wild type과 비교한 결과 다음과 같은 성적을 얻었다. 1. pKSC101을 Eco RI과 Pst I으로 절단하여 940bp insert를 통일 제한효소로 절단한 M 13mp19에 삽입하여 pEP19RF를 구축하였다. 이를 이용하여 dU-SSDNA template를 제조하고, mutagenic primer를 annealing 하여 변이를 도입하였으며, 변이가 도입된 insert를 acceptor plasmid에 삽입시켜 각각 발현 플라스미드 pOEX와 pDEX를 구축하였다. 각각의 mutant 단백질을 발현시키기 위하여 pOEX와 pDEX를 JM109에 형질전환시켜 mutant 재조합 균주인 POMUT109와 DEMUT109를 작성하였다. 2. POMUT109와 DEMUT109를 IPTG 유도 발현시킨 배양상층액을 Vero cell에 대하여 세포독성을 시험한 결과 wild type에 비하여 POMUT109의 배양 여액에서는 2000배, DEMUT109의 배양여액에서는 적어도 3000배 이상의 세포독성을 감소시켰다.

• BMB Reports
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• 제57권2호
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• pp.86-91
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• 2024

The fatty acids content of castor (Ricinus communis L.) seed oil is 80-90% ricinoleic acid, which is a hydroxy fatty acid (HFA). The structures and functional groups of HFAs are different from those of common fatty acids and are useful for various industrial applications. However, castor seeds contain the toxin ricin and an allergenic protein, which limit their cultivation. Accordingly, many researchers are conducting studies to enhance the production of HFAs in Arabidopsis thaliana, a model plant for oil crops. Oleate 12-hydroxylase from castor (RcFAH12), which synthesizes HFA (18:1-OH), was transformed into an Arabidopsis fae1 mutant, resulting in the CL37 line producing a maximum of 17% HFA content. In addition, castor phospholipid:diacylglycerol acyltransferase 1-2 (RcPDAT1-2), which catalyzes the production of triacylglycerol by transferring HFA from phosphatidylcholine to diacylglycerol, was transformed into the CL37 line to develop a P327 line that produces 25% HFA. In this study, we investigated changes in HFA content when endogenous Arabidopsis PDAT1 (AtPDAT1) of the P327 line was edited using the CRISPR/Cas9 technique. The successful mutation resulted in three independent lines with different mutation patterns, which were transmitted until the T4 generation. Fatty acid analysis of the seeds showed that HFA content decreased in all three mutant lines. These findings indicate that AtPDAT1 as well as RcPDAT1-2 in the P327 line are involved in transferring and increasing HFAs to triacylglycerol.

• Toxicological Research
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• 제17권
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• pp.109-115
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• 2001

Abrus agglutinin was purified from the kernels of Abrus precatorius by Sepharose 4B affinity column chromatography followed by Sephadex G-100 gel filtration column chromatography. About 1.25 g of abrus agglutinin was obtained from 1 kg of the kernels. The LD$_{50}$ of abrus agglutinin is 5 mg/kg of body weight, which is less toxic than that of abrin, 20$\mu\textrm{g}$/kg body weight. The amino acid sequence of abrus agglutinin was determined by protein sequencing techniques and deduced from the nucleotide sequence of a cDNA clone encoding full length of abrus agglutinin. There are 258 residues, 2 residues and 267 residues in the A-chain, the linker peptide and the B-chain of abrus agglutinin, respectively. Abrus agglutinin had high homology to abrin-a (77.8%). The 13 amino acid residues involved in catalytic function, which are highly conserved among abrin and ricin, were also conserved within abrus agglutinin. The protein synthesis inhibitory activity of abrus agglutinin ($IC_{50}$/ = 3.5 nM) was weaker than that of abrin-a (0.05 nM). By molecular modeling followed by site-directed mutagenesis showed that Pro199 of abrus agglutinin A-chain located in amphipathic helix H and corresponding to Asn200 of abrin A-chain, can induce bending of helix H. This bending would presumably affect the binding of abrus agglutinin A-chain to its target sequence GpApGpAp, in the tetraloop structure of 285 r-RNA subunit and this could be one of major factors contributing to the relatively weak protein synthesis inhibitory activity and toxicity of abrus agglutinin.n.

• Journal of Microbiology and Biotechnology
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• 제28권5호
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• pp.776-783
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• 2018

The agarase gene gaa16a was identified from a draft genome sequence of Gilvimarinus agarilyticus JEA5, an agar-utilizing marine bacterium. Recently, three agarase-producing bacteria, G. chinensis, G. polysaccharolyticus, and G. agarilyticus, in the genus Gilvimarinus were reported. However, there have been no reports of the molecular characteristics and biochemical properties of these agarases. In this study, we analyzed the molecular characteristics and biochemical properties of agarases in Gilvimarinus. Gaa16A comprised a 1,323-bp open reading frame encoding 441 amino acids. The predicted molecular mass and isoelectric point were 49 kDa and 4.9, respectively. The amino acid sequence of Gaa16A showed features typical of glycosyl hydrolase family 16 (GH16) ${\beta}$-agarases, including a GH16 domain, carbohydrate-binding region (RICIN domain), and signal peptide. Recombinant Gaa16A (excluding the signal peptide and carbohydrate-binding region, rGaa16A) was expressed as a fused protein with maltose-binding protein at its N-terminus in Escherichia coli. rGaa16A had maximum activity at $55^{\circ}C$ and pH 7.0 and 103 U/mg of specific activity in the presence of 2.5 mM $CaCl_2$. The enzyme hydrolyzed agarose to yield neoagarotetraose as the main product. This enzyme may be useful for industrial production of functional neoagaro-oligosaccharides.

• 디지털융복합연구
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• 제16권6호
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• pp.353-361
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• 2018

천연 항균제 및 항산화제 개발의 일환으로 선택한 피마자 열매의 열수 추출물(RFW)과 에탄올 추출물(RFE)은 각각 15.8%와 18.4%의 수율을 나타냈다. Paper-disc method을 통한 항균활성 측정 결과, 피마자 열매를 에탄올로 추출한 추출물에서 항균활성이 나타났다. 특히 녹농균 및 황색포도상구균에서 활성이 우수하였으며, 캔디다균에서도 배양 16시간까지 1.5mm 정도의 항균활성이 나타났으나 24시간 이후에는 캔디다균의 증식이 다시 관찰되었다. 피마자 열매 에탄올추출물(RFE)의 MIC실험 결과 S. aureus와 P. aeruginosa에서 각각 96%와 93%의 항균력이 나타났다. 피마자 잎 열수추출물(RLW)과 피마자 잎 에탄올추출물(RLE)의 DPPH radical 소거능 측정을 통한 항산화 효능 결과는 $1000({\mu}g/m{\ell})$에서 각각 $1.8{\pm}0.6%$, $2.1{\pm}0.7%$의 free radical 소거율을 나타냈다. 본 연구가 향후 천연물을 이용한 항균제 및 항산화제 개발을 위한 기초 자료로 제공되리라 사료된다.