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Enhancement of display efficiency in yeast display system by vector engineering and gene disruption

  • Applied Genetics and Molecular Biotechnology
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Abstract

Vector engineering and gene disruption in host cells were attempted for the enhancement of α-agglutinin-based display of proteins on the cell surface in yeast. To evaluate the display efficiency by flow cytometric analysis, DsRed-monomer fused with FLAG-tag was displayed and immunostained as a model protein. The use of leu2-d in the expression vector resulted in the enhanced efficiency and ratio of the accessible display of proteins. Moreover, the amount of displayed proteins in SED1-disrupted cells increased particularly during the stationary growth phase. The combination of these improvements resulted in the quantitatively enhanced accessible display of DsRed-monomer on the yeast cell surface. The improved yeast display system would be useful in a wider range of its applications in biotechnology.

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References

  • Barberis M, De Gioia L, Ruzzene M, Sarno S, Coccetti P, Fantucci P, Vanoni M, Alberghina L (2005) The yeast cyclin-dependent kinase inhibitor Sic1 and mammalian p27Kip1 are functional homologues with a structurally conserved inhibitory domain. Biochem J 387:639–647

    Article  CAS  Google Scholar 

  • Feldhaus MJ, Siegel RW, Opresko LK, Coleman JR, Feldhous JM, Yeung YA, Cochran JR, Heinzelman P, Colby D, Swers J, Graff C, Wiley HS, Wittrup KD (2003) Flow-cytometric isolation of human antibodies from a nonimmune Saccharomyces cerevisiae surface display library. Nat Biotechnol 21:163–170

    Article  CAS  Google Scholar 

  • Hopp TP, Prickett KS, Price VL, Libby RT, March CJ, Cerretti DP, Urdal DL, Conlon PJ (1988) A short polypeptide marker sequence useful for recombinant protein identification and purification. Bio/Technology 6:1204–1210

    Article  CAS  Google Scholar 

  • Kapteyn JC, Montijn RC, Vink E, de la Cruz J, Llobell A, Douwes JE, Lipke PN, Klis FM (1996) Retention of Saccharomyces cerevisiae cell wall proteins through a phosphodiester-linked β-1,3-/β-1,6-glucan heteropolymer. Glycobiology 6:337–345

    Article  CAS  Google Scholar 

  • Kondo A, Ueda M (2004) Yeast cell-surface display—applications of molecular display. Appl Micobiol Biotechnol 64:28–40

    Article  CAS  Google Scholar 

  • Kuroda K, Shibasaki S, Ueda M, Tanaka A (2001) Cell surface-engineered yeast displaying a histidine oligopeptide (hexa-His) has enhanced adsorption of and tolerance to heavy metal ions. Appl Microbiol Biotechnol 57:697–701

    Article  CAS  Google Scholar 

  • Lee SY, Choi JH, Xu A (2003) Microbial cell-surface display. Trends Biotechnol 21:45–52

    Article  CAS  Google Scholar 

  • Lin Y, Tsumuraya T, Wakabayashi T, Shiraga S, Fujii I, Kondo A, Ueda M (2003) Display of a functional hetero-oligomeric catalytic antibody on the yeast cell surface. Appl Microbiol Biotechnol 62:226–232

    Article  CAS  Google Scholar 

  • Macreadie IG, Horaitis O, Verkuylen AJ, Savin KW (1991) Improved shuttle vectors for cloning and high-level Cu2+-mediated expression of foreign genes in yeast. Gene 104:107–111

    Article  CAS  Google Scholar 

  • Shimoi H, Kitagaki H, Ohmori H, Imura Y, Ito K (1998) Sed1p is a major cell wall protein of Saccharomyces cerevisiae in the stationary phase and is involved in lytic enzyme resistance. J Bacteriol 180:3381–3387

    Article  CAS  Google Scholar 

  • Sikorski RS, Hieter P (1989) A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. Genetics 122:19–27

    CAS  PubMed  PubMed Central  Google Scholar 

  • Takahashi S, Ueda M, Tanaka A (2001) Function of the prosequence for in vivo folding and secretion of active Rhizopus oryzae lipase in Saccharomyces cerevisiae. Appl Microbiol Biotechnol 55:454–462

    Article  CAS  Google Scholar 

  • Takayama K, Suye S, Kuroda K, Ueda M, Kitaguchi T, Tsuchiyama K, Fukuda T, Chen W, Mulchandani A (2006) Surface display of organophosphorus hydrolase on Saccharomyces cerevisiae. Biotechnol Prog 22:939–943

    Article  CAS  Google Scholar 

  • Ueda M (2004) Future direction of molecular display by yeast-cell surface engineering. J Mol Catal B Enzym 28:139–143

    Article  CAS  Google Scholar 

  • Voss S, Skerra A (1997) Mutagenesis of a flexible loop in streptavidin leads to higher affinity for the Strap-tag II peptide and improved performance in recombinant protein purification. Protein Eng 10:975–982

    Article  CAS  Google Scholar 

  • Washida M, Takahashi S, Ueda M, Tanaka A (2001) Spacer-mediated display of active lipase on the yeast cell surface. Appl Microbiol Biotechnol 56:681–686

    Article  CAS  Google Scholar 

  • Ye K, Shibasaki S, Ueda M, Murai T, Kamasawa N, Osumi M, Shimizu K, Tanaka A (2000) Construction of an engineered yeast with glucose-inducible emission of green fluorescence from the cell surface. Appl Microbiol Biotechnol 54:90–96

    Article  CAS  Google Scholar 

  • Zlotnik H, Fernandez MP, Bowers B, Cabib E (1984) Saccharomyces cerevisiae mannoproteins form an external cell wall layer that determines wall porosity. J Bacteriol 159:1018–1026

    Article  CAS  Google Scholar 

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Authors and Affiliations

  1. Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-oiwake-cho, Sakyo-ku, Kyoto, 606-8502, Japan

    Kouichi Kuroda, Ken Matsui, Shinsuke Higuchi & Mitsuyoshi Ueda

  2. Research Institute, Gekkeikan Co. Ltd., 101 Shimotoba-koyanagi-cho, Fushimi-ku, Kyoto, 612-8385, Japan

    Atsushi Kotaka, Hiroshi Sahara & Yoji Hata

Authors
  1. Kouichi Kuroda
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  2. Ken Matsui
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  3. Shinsuke Higuchi
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  4. Atsushi Kotaka
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  5. Hiroshi Sahara
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  6. Yoji Hata
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  7. Mitsuyoshi Ueda
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Correspondence to Mitsuyoshi Ueda.

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Kuroda, K., Matsui, K., Higuchi, S. et al. Enhancement of display efficiency in yeast display system by vector engineering and gene disruption. Appl Microbiol Biotechnol 82, 713–719 (2009). https://doi.org/10.1007/s00253-008-1808-4

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  • DOI: https://doi.org/10.1007/s00253-008-1808-4

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