Effect of Nucleotide Sequences of Polypeptide Linkers on Production of Soluble Single-Chain Fv Antibodies

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Effects of nucleotide sequences of polypeptide linkers on production of soluble single-chain Fv antibodies in <I>E. coli</I> were studied using scFv genes, in which genes of V<SUB>H</SUB> and V<SUB>L</SUB> domains were genetically linked with 4 types of linker genes with or without rare codons, namely genes of scFv (G<SUB>4</SUB>S)<SUB>3</SUB>, scFv (G<SUB>4</SUB>S)<SUB>3</SUB><SUP>R</SUP>, scFv No. 10 and scFv No. 10<SUP>NR</SUP>. The transformants expressing scFvs having linkers with 3–4 rare codons (scFv (G<SUB>4</SUB>S)<SUB>3</SUB><SUP>R</SUP> and scFv No. 10) could grow to higher cell concentrations under induction condition, while growth of the cells expressing scFvs having linkers without rare codon (scFv (G<SUB>4</SUB>S)<SUB>3</SUB> and scFv No. 10<SUP>NR</SUP>) stopped after 3 h. Productivity levels of soluble scFv by cells expressing scFv (G<SUB>4</SUB>S)<SUB>3</SUB><SUP>R</SUP> and scFv No. 10 were higher than those of scFv (G<SUB>4</SUB>S)<SUB>3</SUB> and scFv No. 10<SUP>NR</SUP>. The specific productivity levels of inclusion bodies for scFv (G<SUB>4</SUB>S)<SUB>3</SUB><SUP>R</SUP> and scFv No. 10 decreased in comparison with those for scFv (G<SUB>4</SUB>S)<SUB>3</SUB> and scFv No. 10<SUP>NR</SUP>. This might show that decrease in the translation rate at the region coding polypeptide linkers with rare codons has a favorable effect for the subsequent folding process of scFv molecules. Furthermore, from a SPR analysis using an antigen-immobilized sensor chip, scFv (G<SUB>4</SUB>S)<SUB>3</SUB><SUP>R</SUP> showed the same binding affinity as the original scFv (G<SUB>4</SUB>S)<SUB>3</SUB>. Thus, nucleotide sequences of polypeptide linkers are important to increase productivity of scFv expressed in soluble fraction. The nucleotide sequence of the flexible linker (G<SUB>4</SUB>S)<SUB>3</SUB><SUP>R</SUP> used in this study may be useful for efficient production of various scFvs without loss of the binding affinity of target scFv.

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  • Journal of chemical engineering of Japan  

    Journal of chemical engineering of Japan 41(5), 413-419, 2008-05-01 

    The Society of Chemical Engineers, Japan

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各種コード

  • NII論文ID(NAID)
    10021110329
  • NII書誌ID(NCID)
    AA00709658
  • 本文言語コード
    ENG
  • 資料種別
    ART
  • ISSN
    00219592
  • NDL 記事登録ID
    9498335
  • NDL 雑誌分類
    ZP1(科学技術--化学・化学工業)
  • NDL 請求記号
    Z53-R395
  • データ提供元
    CJP書誌  CJP引用  NDL  J-STAGE 
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