• Title/Summary/Keyword: Molecular characterization

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Characterization of Caveola-Vesicle Complexes (CVCs) Protein, PHIST/CVC-8195 in Plasmodium vivax

  • Wang, Bo;Lu, Feng;Han, Jin-Hee;Lee, Seong-Kyun;Cheng, Yang;Nyunt, Myat Htut;Ha, Kwon-Soo;Hong, Seok-Ho;Park, Won Sun;Han, Eun-Taek
    • Parasites, Hosts and Diseases
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    • v.54 no.6
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    • pp.725-732
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    • 2016
  • Plasmodium vivax produces numerous caveola-vesicle complex (CVC) structures beneath the membrane of infected erythrocytes. Recently, a member helical interspersed subtelomeric (PHIST) superfamily protein, $PcyPHIST/CVC-81_{95}$, was identified as CVCs-associated protein in Plasmodium cynomolgi and essential for survival of this parasite. Very little information has been documented to date about $PHIST/CVC-81_{95}$ protein in P. vivax. In this study, the recombinant $PvPHIST/CVC-81_{95}$ N and C termini were expressed, and immunoreactivity was assessed using confirmed vivax malaria patients sera by protein microarray. The subcellular localization of $PvPHIST/CVC-81_{95}$ N and C termini in blood stage parasites was also determined. The antigenicity of recombinant $PvPHIST/CVC-81_{95}$ N and C terminal proteins were analyzed by using serum samples from the Republic of Korea. The results showed that immunoreactivities to these proteins had 61% and 43% sensitivity and 96.9% and 93.8% specificity, respectively. The N terminal of $PvPHIST/CVC-81_{95}$ which contains transmembrane domain and export motif (PEXEL; RxLxE/Q/D) produced CVCs location throughout the erythrocytic-stage parasites. However, no fluorescence was detected with antibodies against C terminal fragment of $PvPHIST/CVC-81_{95}$. These results suggest that the $PvPHIST/CVC-81_{95}$ is localized on the CVCs and may be immunogenic in natural infection of P. vivax.

Cloning and Characterization of a Novel Carboxylesterase Gene from Cow Rumen Metagenomic Library (소 반추위 메타게놈에서 새로운 carboxylesterase 유전자 클로닝 및 유전산물의 특성)

  • Asraful Islam, Shah Md.;Kim, Min-Keun;Renukaradhya, K. Math;Srinivasa, Reddy R.N.;Kim, Eun-Jin;Kim, Jung-Ho;Kim, Hoon;Yun, Han-Dae
    • Journal of Life Science
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    • v.20 no.9
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    • pp.1306-1313
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    • 2010
  • The gene encoding esterase enzyme was cloned from a metagenomic library of cow rumen bacteria. The esterase gene (est1R) was 2,465 bp in length, encoding a protein of 366 amino acid residues, and the molecular weight of the enzyme was 61,166 Da. Est1R of rumen cosmid library shared 5.9% amino acid identity with Est1R (P37967) of PNB carboxylesterase, 6.1% with Est1R (1EEAA) of acetylcholinesterase and 6.1% with Est1R (1H23A) of chain A. BlastP in NCBI database analysis of Est1R revealed that it was not homologous to previous known lipases and esterases. Est1R showed optimum activity at pH 7.0 and $40^{\circ}C$. On the other hand, the enzyme was found to be most active without organic solvent, followed by 95% activity with methanol, and the enzyme activity was highly affected by hexane (lost 51% activity). Therefore, the novel esterase gene est1R is likely obtainable from cow rumen metagenome and may be utilized for industrial purposes.

Epidemiological Characterization of Adenovirus and Human Bocavirus Detected Acute Respiratory Patients in Busan (부산지역 호흡기감염증 환자로부터 분리한 아데노바이러스와 보카바이러스의 유행양상 분석)

  • Hwang, Su-Jeong;Kim, Nam-Ho;Park, Dong-Ju;Ku, Pyung-Tae;Lee, Mi-Ok;Jin, Sung-Hyun
    • Journal of Life Science
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    • v.27 no.3
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    • pp.275-282
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    • 2017
  • Adenovirus (ADV) and human bocavirus (hBoV) cause acute respiratory tract infections, and are often associated with increased rates of hospitalization and death, particularly in infants and young children. The aim of this study was to analyze the clinical features and molecular phylogeny of ADV and hBoV isolated in Busan, from January 2011 to November 2013. In total, 3,230 specimens (throat swabs) were collected from patients with influenza-like illnesses and acute respiratory tract infections. Multiplex real-time RT-PCR was performed to detect eight respiratory viru [rhinovirus, adenovirus, respiratory syncytial virus, human coronavirus, human metapneumovirus, human bocavirus, parainfluenza virus and influenza virus] and detected 1,485(46.0%) cases. Among 1,485 positive specimens, 257(8.0%) cases were ADV and 68(2.1%) cases of hBoV. A significant clinical feature of ADV is fever and headache whereas hBoV is wheezing. Serotypic distributions of isolated ADV and hBoV were analyzed by sequencing of hexon and VP1/VP2 gene, respectively. ADV was identified seven different serotypes(1~6, 8), revealing a high similarity among the isolates (>97%). The predominant types of ADV were type 1 in 2011, type 3 and 4 in 2012, type 3 in 2013, respectively. ADV type 3 was major causative type during outbreaks in 2013. All of the hBoV was identified as hBoV type 1.

Synthesis and Characterization of Power Conversion Efficiency of D/A Structure Conjugated Polymer Based on Benzothiadiazole-Benzodithiophene (Benzothiadiazole-benzodithiophene을 기반으로 한 D/A구조의 공액 고분자 합성 및 광전변환 효율 특성 개선 연구)

  • Seong, Ki-Ho;Yun, Dae-Hee;Woo, Je-Wan
    • Applied Chemistry for Engineering
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    • v.24 no.5
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    • pp.537-543
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    • 2013
  • In this study, the push-pull structure polymer for organic photo voHaics (OPVs) was synthesized and characterized. The poly{4,8-didodecyloxybenzo[1,2-b;3,4-b]dithiophene-alt-5,6-bis(octyloxy)-4,7-di(thiophen-2-yl)benzo[c][1,2,5]-thiadiazole} (PDBDT-TBTD) was synthesized by Stille coupling reaction using the benzothiadiazole (BTD) derivative as an electron acceptor and benzodithiophene (BDT) derivative as an electron donor. The structure of monomers and polymers was identified by $^1H-NMR$ and GC-MS. The optical, physical and electrochemical properties of the conjugated polymer were identified by GPC, TGA, UV-Vis and cyclic voltammetry. The number average molecular weight ($M_n$) and initial decomposition temperature (5% weight loss temperature, $T_d$) of PDBDT-TBTD were 6200 and $323^{\circ}C$, respectively. The absorption maxima on the film was about 599 nm and the optical band gap was about 1.70 eV. The structure of device was ITO/PEDOT : PSS/PDBDT-TBTD : $PC_{71}BM/BaF_2/Ba/Al$. PDBDT-TBTD and $PC_{71}BM$ were blended with the weight ratio of 1:2 which were then used as an optical active layer. The power conversion efficiency (PCE) of fabricated device was measured by solar simulator and the best PCE was 2.1%.

Progress of Nanofiltration Hollow Fiber Membrane (NF용 중공사 분리막의 발전)

  • Jang, Hanna;Kim, Seongjoong;Lee, Yongtaek;Lee, Kew-Ho
    • Applied Chemistry for Engineering
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    • v.24 no.5
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    • pp.456-470
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    • 2013
  • Hollow-fiber membranes, is one of the new technologies that is growing rapidly in the past few decades. In addition, separation membranes using polymer materials, have attracted attentions in various fields including gas separation, fuel cells, water treatment, wastewater treatment, and organic separation. Nanofiltration (NF) membranes having the separation characteristics in the intermediate range between ultrafiltration and reverse osmosis (RO) membranes for liquid separation, with relatively low investment cost and operating pressure lower than that of RO membranes, have high permeance and rejection performance of multivalent ions as well as organic compounds of molecular weight between $200{\sim}1000gmol^{-1}$. In this paper, we would like to review the research trends on the various structure control and characterization of NF hollow fiber membranes with respect to materials and the methods of preparation (phase inversion method and interfacial polymerization method). Currently, most of NF membranes have been manufactured by plate and frame types or spiral wound types. But hollow fiber types have delayed in commercial products, because of the weak strength when to produce on the basis of the existing materials, therefore the development of new materials or improvement of existing materials will be needed. If improving manufacturing technology is available, hollow fiber types will replace spiral wound types and gradually show a higher market share.

Purification and Characterization of Inulinase from Penicillium sp. (Penicillium sp. 유래 Inulinase의 정제 및 특성)

  • Seok-Yong Kim;Seok-Jong Suh;Seon-Hwa Ha;Seon-Kap Hwang;Joo-Hyun Nam;Dong-Sun Lee;Soon-Duck Hong;Jong-Guk Kim
    • Journal of Life Science
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    • v.8 no.5
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    • pp.614-621
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    • 1998
  • An extracellular inulinase from Penicillium sp. which isolated from soil sample was purified to a single protein th-rough ammonium sulfate fractionation, DEAE-Sephacel ion exchange chromatography and Toyopearl HW 65 F gel filtration. The temperature and pH for the enzyme reaction were around 6$0^{\circ}C$ and 4.0, respectively. The enzyme was stable at 3$0^{\circ}C$-5$0^{\circ}C$ and in the pH range of 4 to 5. $CuCl_2$, $HgCl_2$ and EDTA inhibited the enzyme activity strongly. By contrast, $MnCl_2$ and $CaCl_2$ activated the enzyme activity. The molecular weight of the purified enzyme was esti-mated to be 77,000 dalton by SDS-polyacrylamide gel electrophoresis. The Km values of the enzyme for inulin were calculated to be $2.2\times10^{-3}$M. TLC analysis suggested that purified enzyme is exo-type inulinase. The NH2-terminal amino acid sequences of the purified enzyme was determined to be $NH_2$-X-Glu-Ser-Tyr-Thr-Glu-Lys/Leu-Tyr-Arg-Pro.

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Characterization and Expression of Penaeidin 3-2 from Fleshy Prawn Fenneropenaeus chinensis (대하 Penaeidin 3-2 유전자의 동정 및 발현)

  • Park, Eun-Mi;Cho, Hyun Kook;Hong, Gyeong-Eun;Nam, Bo-Hye;Kim, Young-Ok;Kim, Woo-Jin;Lee, Sang-Jun;Han, Hyon Sob;Lee, Jae Yong;Kim, Jong-Sheek;Jang, In-Kwon;Cheong, JaeHun;Choi, Tae-Jin;Kong, Hee Jeong
    • Journal of Marine Bioscience and Biotechnology
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    • v.2 no.1
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    • pp.34-39
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    • 2007
  • Penaeidins are members of a special family of antimicrobial peptides existing in several species of shrimp and play a crucial role in the immunological defense of shrimp. In this study, we isolated and characterized one EST clone (penaeidin) from cDNA library of fleshy prawn Fenneropenaeus chinensis hemocytes. Amino acids sequence comparison and phylogenetic analysis with other known penaeidins revealed that our clone was completely identical to F. chinensis Penaeidin 3-2 (Accession no. ABC33920), which composed of 71 amino acids with a putative signal peptide (1-19) and a cysteine-rich domain (C-terminal part). The expression and distribution of Penaeidin 3-2 transcripts in shrimp were detected in hemocytes, hepatopancreas, and muscles, and that Penaeidin 3-2 was constitutively expressed mainly in hemocytes. The artificial infection of white spot syndrome virus to F. chinensis resulted in Penaeidin 3-2 mRNA up-regulation in hemocytes, suggesting that the possible role of Penaeidin 3-2 in host defense system of F. chinensis.

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Isolation and Characterization of Bacillus licheniformis SC082 Degrading Fibrin and Chitin from Shrimp Jeot-Gal (새우젓으로부터 혈전과 chitin 분해능을 지닌 균주 Bacillus licheniformis SC082의 분리 및 특성)

  • Cho, Eun-Kyung;Jung, Yu-Jung;Gal, Sang-Wan;Choi, Young-Ju
    • Journal of Life Science
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    • v.19 no.10
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    • pp.1424-1431
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    • 2009
  • Shrimp Jeot-Gal is a popular traditional Korean fermented seafood and has been used for seasoning. We isolated a bacterium showing strong extra-cellular fibrinolysis and chitinase activity from shrimp Jeot-Gal and the strain was designated SC082. SC082 was identified as Bacillus licheniformis by 16S rRNA sequence homology search. B. licheniformis SC082 exhibited optimum temperature, pH, and salt concentration at $37^{\circ}C$, pH 7.0, and 6%, respectively. Substrate specificity of the culture supernatant from B. licheniformis SC082 was detected in fibrin, skim milk, and chitin plate. The fibrinolytic activity was highly maintained up to $50^{\circ}C$ at a pH of 7.0 for 3 hr and was stable up to pH 9.0 at $37^{\circ}C$ for 3 hr. The chitinase activity was remarkably induced by addition of 1.0% colloidal chitin and the pH and temperature optima of the enzyme were 5.0 and $45^{\circ}C$, respectively. In sodium dodecyl sulfate-polyacrylamide gel electrophoresis and zymogram analysis, this strain produced three fibrinolytic isozymes and two chitinase isozymes. The approximate molecular weights of the putative fibrinolytic enzymes were 23.0, 62.0, and 72.0 kDa and those of the chitinases were 62.0 and 55.0 kDa, respectively. The antioxidant activity of SC082 was also measured by using 2,2-diphenyl-l-picryl-hydrazyl (DPPH) free radical. The DPPH radical scavenging was slightly increased in a dose-dependent manner.

Purification and Characterization of Mitochondrial Malate Dehydrogenase during Ovarian Development in Aedes aegypti L. (Aedes aegypti L. 난성숙과정중 생성되는 Mitochondrial Malate Dehydrogenase의 정제 및 특성)

  • 김인규;이강석;정규회;박영민;성기창
    • Korean journal of applied entomology
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    • v.34 no.3
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    • pp.181-190
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    • 1995
  • Malate dehydrogenase in the mosquito ovary after a blood meal, Aedes aegypti, was purified and characterized. MDH purification steps involved DEAE-Sepharose, S-Sepharose and Cibacron blue affinity chromatography. The purified MDH was 70,000 daltons in molecular weight and was a homodimer consisting of tow identical subunits. Optimal activity of purified MDH was obtained pH 9.0-9.2 in malate-oxaloacetate reaction and pH 9.8-10.2, in oxaloactate-malate reaction. With obtained pH 9.0-92 in malate-oxaloacetate reaction and pH 9.8-10.2, in oxaloactate-malate reaction. With malate as substrate, purified mitochondrial MDH (1.28$\times$${10}^{-4}$ M) had lower Km value than cytoplasmic MDH (8.92x${10}^{-3}$ M). MDH activity was inhibited by citrate, $\alpha$-ketoglutarate, and ATP. Inhibition of MDH activity by ATP and citrate was less in malate-oxaloacetate reaction and in oxaloacetate-malate reaction. MDH activity was completely inhibited by ATP in oxaloacetate-malate reaction and not inhibited by citrate in malate-oxaloacetate reaction. Temporal activity change of MDH is similar to that of isocitrate dehydrogenase in the ovary after blood feeding; their activities in the ovary began to rise at 18 hours after a blood meal, and reached at the maximal level at 48 hours.

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Molecular characterization and docking dynamics simulation prediction of cytosolic OASTL switch cysteine and mimosine expression in Leucaena leucocephala

  • Harun-Ur-Rashid, Md.;Masakazu, Fukuta;Amzad Hossain, Md.;Oku, Hirosuke;Iwasaki, Hironori;Oogai, Shigeki;Anai, Toyoaki
    • Proceedings of the Korean Society of Crop Science Conference
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    • 2017.06a
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    • pp.36-36
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    • 2017
  • Out of twenty common protein amino acids, there are many kinds of non protein amino acids (NPAAs) that exist as secondary metabolites and exert ecological functions in plants. Mimosine (Mim), one of those NPAAs derived from L. leucocephala acts as an iron chelator and reversely block mammalian cell cycle at G1/S phases. Cysteine (Cys) is decisive for protein and glutathione that acts as an indispensable sulfur grantor for methionine and many other sulfur-containing secondary products. Cys biosynthesis includes consecutive two steps using two enzymes-serine acetyl transferase (SAT) and O-acetylserine (thiol)lyase (OASTL) and appeared in plant cytosol, chloroplast, and mitochondria. In the first step, the acetylation of the ${\beta}$-hydroxyl of L-serine by acetyl-CoA in the existence of SAT and finally, OASTL triggers ${\alpha}$, ${\beta}$-elimination of acetate from OAS and bind $H_2S$ to catalyze the synthesis of Cys. Mimosine synthase, one of the isozymes of the OASTLs, is able to synthesize Mim with 3-hydroxy-4-pyridone (3H4P) instead of $H_2S$ for Cys in the last step. Thus, the aim of this study was to clone and characterize the cytosolic (Cy) OASTL gene from L. leucocephala, express the recombinant OASTL in Escherichia coli, purify it, do enzyme kinetic analysis, perform docking dynamics simulation analysis between the receptor and the ligands and compare its performance between Cys and Mim synthesis. Cy-OASTL was obtained through both directional degenerate primers corresponding to conserved amino acid region among plant Cys synthase family and the purified protein was 34.3KDa. After cleaving the GST-tag, Cy-OASTL was observed to form mimosine with 3H4P and OAS. The optimum Cys and Mim reaction pH and temperature were 7.5 and $40^{\circ}C$, and 8.0 and $35^{\circ}C$ respectively. Michaelis constant (Km) values of OAS from Cys were higher than the OAS from Mim. Inter fragment interaction energy (IFIE) of substrate OAS-Cy-OASTL complex model showed that Lys, Thr81, Thr77 and Gln150 demonstrated higher attraction force for Cys but 3H4P-mimosine synthase-OAS intermediate complex showed that Gly230, Tyr227, Ala231, Gly228 and Gly232 might provide higher attraction energy for the Mim. It may be concluded that Cy-OASTL demonstrates a dual role in biosynthesis both Cys and Mim and extending the knowledge on the biochemical regulatory mechanism of mimosine and cysteine.

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