• Title/Summary/Keyword: library screening

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Identification of Small GTPases That Phosphorylate IRF3 through TBK1 Activation Using an Active Mutant Library Screen

  • Jae-Hyun Yu;Eun-Yi Moon;Jiyoon Kim;Ja Hyun Koo
    • Biomolecules & Therapeutics
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    • v.31 no.1
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    • pp.48-58
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    • 2023
  • Interferon regulatory factor 3 (IRF3) integrates both immunological and non-immunological inputs to control cell survival and death. Small GTPases are versatile functional switches that lie on the very upstream in signal transduction pathways, of which duration of activation is very transient. The large number of homologous proteins and the requirement for site-directed mutagenesis have hindered attempts to investigate the link between small GTPases and IRF3. Here, we constructed a constitutively active mutant expression library for small GTPase expression using Gibson assembly cloning. Small-scale screening identified multiple GTPases capable of promoting IRF3 phosphorylation. Intriguingly, 27 of 152 GTPases, including ARF1, RHEB, RHEBL1, and RAN, were found to increase IRF3 phosphorylation. Unbiased screening enabled us to investigate the sequence-activity relationship between the GTPases and IRF3. We found that the regulation of IRF3 by small GTPases was dependent on TBK1. Our work reveals the significant contribution of GTPases in IRF3 signaling and the potential role of IRF3 in GTPase function, providing a novel therapeutic approach against diseases with GTPase overexpression or active mutations, such as cancer.

Isolation of Novel Pseudonocardia Polyene Biosynthetic Genes via Genomics-based PCR Screening

  • Lee, Mi-Yeon;Hwang, Young-Bin;Park, Hyun-Joo;Han, Kyu-Boem;Kim, Eung-Soo
    • 한국생물공학회:학술대회논문집
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    • 2005.04a
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    • pp.396-397
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    • 2005
  • The polyene antibiotics are a family of most promising antifungal polyketide compounds, typically produced by actinomycetes species. Using the polyene CYP-specific PCR screening with served actinomycetes genomic DNAs, Pseudonocardia autotrophica strain was identified to contain a unique polyene-specific CYP gene. The genomic DNA library screening using the polyene-specific CYP gene probe revealed the positive cosmid clone containing an approximately 34.5 kb DNA fragment revealed a total of seven complete and two incomplete open reading frame (ORFs), which are highly homologous but unique to previously-known polyene biosynthetic genes. These results suggest that the polyene-specific screening approach should be an efficient way of isolating potectially-valuable cryptic polyene biosynthetic gene cluster from various rare actinomycetes.

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Study on the development process and operations of aviation security screening (항공 보안검색의 발전과정과 운영실태에 관한 연구)

  • Kim, Yong-Wook
    • Korean Security Journal
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    • no.7
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    • pp.61-93
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    • 2004
  • The terrorism threat against aviation industry is increasing after 9/11 incidents. As the way of preventing the future threat, this study is to review current status and make practical suggestions on Incheon International Airport security screening. The Library research method is utilized for this study. Chapter 1 is an introduction part and describes the general instruction. Chapter 2 describes the importance of aviation security and cases of aviation terrorism. Screening history is described in chapter 3 detailing operations and equipment. Status of IIAC(Incheon International Airport Corporation) security screening and prevention measures are described in chapter 4 and chapter 5 describes the final conclusion. In conclusion, if the introduction of state-of-the-art security equipment and modification of inadequate regulations come in first, whatever the threat exists, aviation terrorism can be prevented with the positive prevention efforts.

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Cyclic Peptides as Therapeutic Agents and Biochemical Tools

  • Joo, Sang-Hoon
    • Biomolecules & Therapeutics
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    • v.20 no.1
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    • pp.19-26
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    • 2012
  • There are many cyclic peptides with diverse biological activities, such as antibacterial activity, immunosuppressive activity, and anti-tumor activity, and so on. Encouraged by natural cyclic peptides with biological activity, efforts have been made to develop cyclic peptides with both genetic and synthetic methods. The genetic methods include phage display, intein-based cyclic peptides, and mRNA display. The synthetic methods involve individual synthesis, parallel synthesis, as well as split-and-pool synthesis. Recent development of cyclic peptide library based on split-and-pool synthesis allows on-bead screening, in-solution screening, and microarray screening of cyclic peptides for biological activity. Cyclic peptides will be useful as receptor agonist/antagonist, RNA binding molecule, enzyme inhibitor and so on, and more cyclic peptides will emerge as therapeutic agents and biochemical tools.

Solution-Phase Strategies for the Design, Synthesis, and Screening of Libraries Based on Natural Products

  • Kim, Sang-Hee
    • Proceedings of the PSK Conference
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    • 2003.10a
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    • pp.88-88
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    • 2003
  • The syntheses of different types of stilbenoid libraries have been studied recently. In these courses, the screening of the generated natural product-mimic focused libraries led to the identification of the novel lead compounds for human cytochrome P450 (CYP) lAs, melanin production, and sortase A. A library of trans-stilbene derivatives was prepared through a new efficient solution pahse synthetic pathway and their inhibitory activities were evaluated on human cytochrome P450s(CYP) 1A1, 1A2, and 1B1 to find a potent and selective CYP1 inhibitor. (omitted)

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Design of Novel Ras Farnesyltransferase Inhibitors Based on Virtual Screening and Docking Studies

  • Jung, Kang-Rae;Park, Hyung-Yeon;Kim, Chan-Kyung;Lee, Bon-Su
    • Proceedings of the PSK Conference
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    • 2003.10b
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    • pp.175.2-175.2
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    • 2003
  • Inhibition of the protein-modifying enzyme farnesyltransferase is considered as a major emerging strategy in cancer therapy because of the involvement of farnesylated proteins in oncogensis. We studied the structure-activity relationship of a novel class of CAAX-peptidomimetic farnesyltransferase inhibitors based on the benzophenone scaffold. FlexX docking of inhibitors confirmed reasonable fit of the molecule into the peptide binding site of farnesyltransferase. We also performed a virtual screening with LeadQuest chemical library databases to idenfity novel inhibitors of farnesyltransferase. (omitted)

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Lipase Diversity in Glacier Soil Based on Analysis of Metagenomic DNA Fragments and Cell Culture

  • Zhang, Yuhong;Shi, Pengjun;Liu, Wanli;Meng, Kun;Bai, Yingguo;Wang, Guozeng;Zhan, Zhichun;Yao, Bin
    • Journal of Microbiology and Biotechnology
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    • v.19 no.9
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    • pp.888-897
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    • 2009
  • Lipase diversity in glacier soil was assessed by culture-independent metagenomic DNA fragment screening and confirmed by cell culture experiments. A set of degenerate PCR primers specific for lipases of the hormone-sensitive lipase family was designed based on conserved motifs and used to directly PCR amplify metagenomic DNA from glacier soil. These products were used to construct a lipase fragment clone library. Among the 300 clones sequenced for the analysis, 201 clones encoding partiallipases shared 51-82% identity to known lipases in GenBank. Based on a phylogenetic analysis, five divergent clusters were established, one of which may represent a previously unidentified lipase subfamily. In the culture study, 11 lipase-producing bacteria were selectively isolated and characterized by 16S rDNA sequences. Using the above-mentioned degenerate primers, seven lipase gene fragments were cloned, but not all of them could be accounted for by the clones in the library. Two full-length lipase genes obtained by TAIL-PCR were expressed in Pichia pastoris and characterized. Both were authentic lipases with optimum temperatures of ${\le}40^{\circ}C$. Our study indicates the abundant lipase diversity in glacier soil as well as the feasibility of sequence-based screening in discovering new lipase genes from complex environmental samples.

A study on the screening of toxic materials by HPTLC and GC/MS (HPTLC 및 GC/MS를 이용한 유해화학물질의 스크리닝에 관한 연구)

  • Park, Sung-Woo;Jang, Seong-Gil;Park, You-Sin;Lee, Jin-Hoon;Lee, Sang-Ki;You, Jae-Hoon;Kim, Dong-Hwan;Jin, Kwang-Ho;Kim, Ki-Wook;Kim, Yu-Na;Lho, Dong-Seok
    • Analytical Science and Technology
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    • v.13 no.1
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    • pp.108-120
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    • 2000
  • To perform an effective screening for toxic materials of forensic interest detected in high profile criminal case in biological and environmental samples, we tried to construct a searchable computerized database using HPTLC(High Performance Thin Layer Chromatography) and GC/MS. Retardation factor($R_f$) values and UV spectral data of HPTLC were investigated for 160 pesticides, 34 chemicals and 39 explosives of standard grade. The data were compiled in a library. We also analyzed 112 pesticides, 31 chemicals and 17 explosives and 57 volatile organic compounds(VOCs) by GC/MS. The data for RT and characteristic mass ions were also compiled in a library.

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