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Chemogenomics Profiling of Drug Targets of Peptidoglycan Biosynthesis Pathway in Leptospira interrogans by Virtual Screening Approaches

  • Bhattacharjee, Biplab;Simon, Rose Mary;Gangadharaiah, Chaithra;Karunakar, Prashantha
    • Journal of Microbiology and Biotechnology
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    • v.23 no.6
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    • pp.779-784
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    • 2013
  • Leptospirosis is a worldwide zoonosis of global concern caused by Leptospira interrogans. The availability of ligand libraries has facilitated the search for novel drug targets using chemogenomics approaches, compared with the traditional method of drug discovery, which is time consuming and yields few leads with little intracellular information for guiding target selection. Recent subtractive genomics studies have revealed the putative drug targets in peptidoglycan biosynthesis pathways in Leptospira interrogans. Aligand library for the murD ligase enzyme in the peptidoglycan pathway has also been identified. Our approach in this research involves screening of the pre-existing ligand library of murD with related protein family members in the putative drug target assembly in the peptidoglycan biosynthesis pathway. A chemogenomics approach has been implemented here, which involves screening of known ligands of a protein family having analogous domain architecture for identification of leads for existing druggable protein family members. By means of this approach, one murC and one murF inhibitor were identified, providing a platform for developing an anti-leptospirosis drug targeting the peptidoglycan biosynthesis pathway. Given that the peptidoglycan biosynthesis pathway is exclusive to bacteria, the in silico identified mur ligase inhibitors are expected to be broad-spectrum Gram-negative inhibitors if synthesized and tested in in vitro and in vivo assays.

Cloning and functional expression of a cecropin-A gene from the Japanese oak silkworm, Antheraea yamamai (천잠 cecropin-A 유전자 클로닝 및 재조합 발현)

  • Kim, Seong-Ryul;Choi, Kwang-Ho;Kim, Sung-Wan;Goo, Tae-Won;Hwang, Jae-Sam
    • Journal of Sericultural and Entomological Science
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    • v.52 no.1
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    • pp.45-51
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    • 2014
  • A cecropin-A gene was isolated from the immunized larvae of the Japanese oak silkworm, Antheraea yamamai and designed Ay-CecA. The complete Ay-CecA cDNA consists of 419 nucleotides with 195 bp open reading frame encoding a 64 amino acid precursor that contains a putative 22-residue signal peptide, a 4-residue propetide and a 37-residue mature peptide with a theoretical mass of 4046.81. The deduced amino acid sequence of the peptide evidenced a significant degree of identity (62 ~ 78% identity) with other lepidopteran cecropins. Like many insect cecropin, Ay-CecA also harbored a glycine residue for C-terminal amidation at the C-end, which suggests potential amidation. To understand this peptide better, we successfully expressed bioactive recombinant Ay-CecA in Escherichia coli that are highly sensitive to the mature peptide. For this, we fused mature Ay-CecA gene with insoluble protein ketosteroid isomerase (KSI) gene to avoid the cell death during induction. The fusion KSI-CecA protein was expressed as inclusion body. The expressed fusion protein was purified by Ni-NTA immobilized metal affinity chromatography (IMAC), and cleaved by cyanogen bromide (CNBr) to release recombinant Ay-CecA. The purified recombinant Ay-CecA showed considerably antibacterial activity against Gram-negative bacteria, E. cori ML 35, Klebsiella pneumonia and Pseudomonas aeruginosa. Our results proved that this peptide with a potent antibacterial activity may play a role in the immune response of Japanese oak silkworm.

Effects of Heated Effluents on the Intertidal Macroalgal Community Near Gori Nuclear Power Plant (고리원전의 온배수 방출이 주변 해조군집에 미치는 영향)

  • Kim, Young-Hwan;Ahn, Jung-Kwan;Yoon, Hee-Dong;Jang, Min-A
    • ALGAE
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    • v.22 no.4
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    • pp.297-304
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    • 2007
  • This study is intended to clarify the effects of heated effluents on intertidal benthic marine algal community in Korea. The species composition and biomass of marine algae at the discharge canal of Gori nuclear power plant on the southeastern coast of Korea were investigated seasonally from February 2001 to October 2006. As a result, 54 species (7 blue-green, 12 green, 9 brown and 26 red algae) of marine algae were found at the discharge canal during the past six years. In general, the number of species observed was abundant during winter to spring and less in autumn. Enteromorpha compressa, E. intestinalis, E. prolifera and Caulacanthus ustulatus were common species found more than 80% frequency during the study period. Seasonal fluctuations of mean biomass were 1-440 g dry wt m–2 and dominant species in biomass were Enteromorpha spp. (contribution to a total biomass proportion 28%), Sargassum horneri (14%) and Amphiroa beauvoisii (14%). It is evident from the floristic composition and biomass data that unique micro-environment of the discharge canal support different communities from those on the intake or control area. Results from the large numbers of surveys before and during plant operation showed that, in the regions influenced by thermal effluents such as the discharge canal of power plants, the process of ecological succession has been proceeded. It is assumed that the uni-directional water flow and the time of overhaul largely affect the development and succession of benthic marine algal communities of the discharge canal.

The effect of heterogeneous hyperimmune IgG antibody on prophylaxis and treatment of Pneumocystis carinii infection in rats (폐포자충증에 대한 이종항혈청 내 1gG 항체의 예방 및 치료효과)

  • 이미정;조상록
    • Parasites, Hosts and Diseases
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    • v.36 no.2
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    • pp.127-132
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    • 1998
  • Immunotherapy has been used in support of trimethoprim-sulfamethoxazole treatment for Pneumocvstis corinii pneumonia. The present study investigated the therapeutic or preventive effects of heterogeneous hyperimmune IgG antibody (HIA) in experimental rats. Their immunity was suppressed by steroid injection, and they were also injected peritoneally with HIA which reacted with 40-55, 92, 116, and 200 kDa bands of the crude antigen. All rats were infected by p. ccrinii and the cystic forms on lung impression smears were counted. The count was 20.5-76.5 (mean 52.5 ± 19.)1 in those which received steroid only, but decreased to 6.0-21.0 (mean 13.5 : 10.6) in those of group 3 which received HIA for the same duration. In other groups, the mean count ranged from 29.9 t 32.9 to 54.1 t 47.7, and in those which received 13.7 mg HIA the reduction effect was greater than in those which received 6.8 mg or 20.5 mg HIA. The present finding confirmed that in rats during the early stage of infection, the heterogeneous HIA to MSG antigen bands had a partial effect on p. cori,nii pneumonia, both prophylactically and therapeutically.

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Study on the Winter Mass Mortality of red sea bream, Pagrus major in South sea area (동절기 남해안 참돔(Pagrus major)의 대량폐사에 관한 연구)

  • Choi, Hye-Sung;Jung, Sung-Hee;Hur, Young-baek;Yang, Jun-Yong
    • Journal of fish pathology
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    • v.21 no.1
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    • pp.35-43
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    • 2008
  • Pathological symptoms and hematological parameters of red sea bream, Pagrus major and water temperature in the culture ground was investigated to clarlify the cause of winter moratility. Dead fish showed green liver and accumulation of ascites in the cavity. A few Bivagina tai were also found on the gill but either bacteria or virus were not. When hematological parameters from fish taken before/after the winter mortality were compared, blood glucose, serum AST and ALT, and total cholesterol, trigyceride and total protein were significantly decreased in the fish after the winter mortality. These results may explain that the nutritional level of fish was decreased because fish could not fed during winter season. According to the CORI monitoring system operating by KODC, NFRDI a long term water temperature from Dec. 25, 2005 to Feb. 24, 2006 (60 days) were exposed the low water temperature environments to the red sea bream. First of all, mass mortality began at Sarayngdo where low temperature below 8℃ continued for 42 days. The winter mortality did not occured in the depths of 9 m to 19.2 m where difference of water temperature in the surface and bottom was only within 0.1℃. But in the depth of 7.5 m to 11 m winter mortality occured where water temperature in the surface and bottom showed much variation ranged from 0.2℃ to 1.4℃. From these results, great difference of water temperature in the surface and bottom of the culturing area might results in winter mortality of red sea bream.

Immunoblot analysis for serum antibodies to Pneumocystis carinii by age and intensity of infection in rats (흰쥐의 연령과 감염과정에 따른 폐포자충에 대한 항체형성 양상)

  • 홍성태;이미정
    • Parasites, Hosts and Diseases
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    • v.33 no.3
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    • pp.187-194
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    • 1995
  • The present study aims to observe changing patterns of serum antibody to hleumuvstis calinii in normal rats of different ages and in immunosuppressed rats. The serum IgG antibody was observed by immunoblotting with crude antigen of f carinii which were purified from the lungs of infected rats. The crude antigens separated in SDS-PAGE resolved more than 20 protein bands from 20 to 200 kDa. Of them,40-45, 50-55, 116 and 200 kDa bands were major antigens of R cori.nii. Most of the normal rats of up to 4 weeks had the antibodies reacting the 4 bands, but none of 8-week-old rats revealed the specific antibody. After the rats grew for 40 weeks, all were found to have the antibody in their serum. Same pattern of serum antibody level by age was found in ELISA. When immunosuppressed rats became heavily infected, the antibody in their serum decreased distinctively. The present results suggest that antibodies in normal newborn rats are transferred from their mother and lowered up to 8 weeks. Thereafter, the levels of the antibodies begin to increase by natural exposure to R cnrinii. It was also confirmed that the intensity of P cnrinii infection is inversely related with levels of serum antibodies.

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