• Title/Summary/Keyword: Combinatorial library

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Successful Application of the Dual-Vector System II in Creating a Reliable Phage-Displayed Combinatorial Fab Library

  • Song, Suk-yoon;Hur, Byung-ung;Lee, Kyung-woo;Choi, Hyo-jung;Kim, Sung-soo;Kang, Goo;Cha, Sang-hoon
    • Molecules and Cells
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    • v.27 no.3
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    • pp.313-319
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    • 2009
  • The dual-vector system-II (DVS-II), which allows efficient display of Fab antibodies on phage, has been reported previously, but its practical applicability in a phage-displayed antibody library has not been verified. To resolve this issue, we created two small combinatorial human Fab antibody libraries using the DVS-II, and isolation of target-specific antibodies was attempted. Biopanning of one antibody library, termed DVFAB-1L library, which has a $1.3{\times}10^7$ combinatorial antibody complexity, against fluorescein-BSA resulted in successful isolation of human Fab clones specific for the antigen despite the presence of only a single light chain in the library. By using the unique feature of the DVS-II, an antibody library of a larger size, named DVFAB-131L, which has a $1.5{\times}10^9$ combinatorial antibody complexity, was also generated in a rapid manner by combining $1.3{\times}10^7$ heavy chains and 131 light chains and more diverse anti-fluorescein-BSA Fab antibody clones were successfully obtained. Our results demonstrate that the DVS-II can be applied readily in creating phage-displayed antibody libraries with much less effort, and target-specific antibody clones can be isolated reliably via light chain promiscuity of antibody molecules.

Combinatorial Solid Phase Peptide Synthesis and Bioassays

  • Shin, Dong-Sik;Kim, Do-Hyun;Chung, Woo-Jae;Lee, Yoon-Sik
    • BMB Reports
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    • v.38 no.5
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    • pp.517-525
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    • 2005
  • Solid phase peptide synthesis method, which was introduced by Merrifield in 1963, has spawned the concept of combinatorial chemistry. In this review, we summarize the present technologies of solid phase peptide synthesis (SPPS) that are related to combinatorial chemistry. The conventional methods of peptide library synthesis on polymer support are parallel synthesis, split and mix synthesis and reagent mixture synthesis. Combining surface chemistry with the recent technology of microelectronic semiconductor fabrication system, the peptide microarray synthesis methods on a planar solid support are developed, which leads to spatially addressable peptide library. There are two kinds of peptide microarray synthesis methodologies: pre-synthesized peptide immobilization onto a glass or membrane substrate and in situ peptide synthesis by a photolithography or the SPOT method. This review also discusses the application of peptide libraries for high-throughput bioassays, for example, peptide ligand screening for antibody or cell signaling, enzyme substrate and inhibitor screening as well as other applications.

Combinatorial Library and Chemogenomics Approach: Discovery of Protein Secondary Structure Mimetic Small Molecule Inhibitors of Tryptase and Ref-l for Asthma

  • Moon, Sung-Hwan
    • Proceedings of the PSK Conference
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    • 2003.10a
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    • pp.92-92
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    • 2003
  • The drug discovery landscape is changing rapidly in the post-genomic era. Mapping of the human genome has led to an abundance of potential drug targets. Drug discovery times and costs can be significantly reduced by developing methods for high throughput target identification/ validation, multiplexed assay development and high efficient combinatorial chemistry. (omitted)

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High-throughput Preparation and Characterization of Powder and Thin-film Library for Electrode Materials

  • Fujimoto, Kenjiro;Onoda, Kazuhiro;Ito, Shigeru
    • Proceedings of the Korean Powder Metallurgy Institute Conference
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    • 2006.09a
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    • pp.254-255
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    • 2006
  • Powder library of pseudo four components Li-Ni-Co-Ti compounds were prepared for exploring the composition region with the single phase of the layer-type structure by using combinatorial high-throuput preparation system "M-ist Combi" based on electrostatic spray deposition method. The new layer-type compounds were found wider composition region than the previous report. This process is promising way to find multi component functional materials.

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Enhancing the Solubility of Recombinant Akt1 in Escherichia coli with an Artificial Transcription Factor Library

  • Park Kyung-Soon;Lee Ho-Rim;Kim Jin-Soo
    • Journal of Microbiology and Biotechnology
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    • v.16 no.2
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    • pp.299-302
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    • 2006
  • A combinatorial library of artificial transcription factors (ATFs) was introduced into the bacterial cells that expressed the Akt1-GFP fusion protein. By measuring the level of fluorescence generated by the transformed E. coli cells, we were able to obtain clones in which ATFs increased the solubility of the Akt1. Our results show that ATF library is a useful tool for increasing the solubility of selected recombinant proteins in E. coli.

Antibradykinin Effects of the Non-peptide Antagonists of Mixture Libraries Prepared by Solution-phase Combinatorial Synthesis

  • KAM Yoo Lim;CHOO Hea-Young P.
    • Biomolecules & Therapeutics
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    • v.13 no.3
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    • pp.181-184
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    • 2005
  • The solution-phase combinatorial synthesis of iminodiacetic acid triamide libraries linked to 1-(4-chlorobenzhydryl)piperazine has been reported. Ten mixture libraries, each containing 5 components, were synthesized in 4 steps from N-BOC-iminodiacetic acid anhydride. Antibradykinin effects of the mixture and individual libraries were compared using guinea-pig ileum smooth muscle. The changes in the inhibition were also observed by testing the combination of two different compounds from the same library. We found out the correlation between the inhibition of mixtures and that of individual libraries. It is possible to choose the mixtures with relatively high inhibitory effects to find out the most effective individual compound for further synthesis.

Synthesis of combinatorial library of $\beta$-ketoacetoanilide chlorides and their antifungal activity against main plant pathogens ($\beta$-Ketoacetoanilide 염화물의 조합 라이브러리 합성 및 주요 식물병원균에 대한 항균활성)

  • Hahn, Hoh-Gyu;Nam, Kee-Dal;Bae, Su-Yeal;Yang, Bum-Seung;Lee, Seon-Woo;Cho, Kwang-Yun
    • The Korean Journal of Pesticide Science
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    • v.8 no.1
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    • pp.8-15
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    • 2004
  • A synthesis of new $\beta$-ketoacetoanilide chloride derivatives and anti fungal activity of these compounds library against 6 typical plant pathogens were described. Reaction of ketene dimer with chlorine followed by treatment of aniline derivatives gave 89 kinds of the corresponding $\beta$-ketoacetoanilide chlorides through combinatorial synthetic technology using Carousel Reaction Stations. Evaluation of antifungal activity (in vivo) of this chemical library against rice blast, rice sheath blight, tomato aray mold, tomato late blight, wheat leaf rust and barley powdery mildew was carried out. In general, $\beta$-ketoacetoanilide chlorides which present a substituent at 4 in phenyl group(para) of the compounds showed selective control activity against tomato late blight caused by Phytophthora infestans.

Construction of a Large Synthetic Human Fab Antibody Library on Yeast Cell Surface by Optimized Yeast Mating

  • Baek, Du-San;Kim, Yong-Sung
    • Journal of Microbiology and Biotechnology
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    • v.24 no.3
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    • pp.408-420
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    • 2014
  • Yeast surface-displayed antibody libraries provide an efficient and quantitative screening resource for given antigens, but suffer from typically modest library sizes owing to low yeast transformation efficiency. Yeast mating is an attractive method for overcoming the limit of yeast transformation to construct a large, combinatorial antibody library, but the optimal conditions have not been reported. Here, we report a large synthetic human Fab (antigen binding fragment) yeast surface-displayed library generated by stepwise optimization of yeast mating conditions. We first constructed HC (heavy chain) and LC (light chain) libraries, where all of the six CDRs (complementarity-determining regions) of the variable domains were diversified mimicking the human germline antibody repertoires by degenerate codons, onto single frameworks of VH3-23 and $V{\kappa}1$-16 germline sequences, in two haploid cells of opposite mating types. Yeast mating conditions were optimized in the order of cell density, media pH, and cell growth phase, yielding a mating efficiency of ~58% between the two haploid cells carrying HC and LC libraries. We constructed two combinatorial Fab libraries with CDR-H3 of 9 or 11 residues in length with colony diversities of more than $10^9$ by one round of yeast mating between the two haploid HC and LC libraries, with modest diversity sizes of ${\sim}10^7$. The synthetic human Fab yeast-displayed libraries exhibited relative amino acid compositions in each position of the six CDRs that were very similar to those of the designed repertoires, suggesting that they are a promising source for human Fab antibody screening.