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Cell-surface expression of Aspergillus saitoi-derived functional α-1,2-mannosidase on Yarrowia lipolytica for glycan remodeling

  • Systems and Synthetic Microbiology and Bioinformatics
  • Published:
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Abstract

Expression of proteins on the surface of yeast has a wide range of applications, such as development of live vaccines, screening of antibody libraries, and use as whole-cell biocatalysts. The hemiascomycetes yeast Yarrowia lipolytica has been raised as a potential host for heterologous expression of recombinant proteins. In this study, we report the expression of Aspergillus saitoi α-1,2-mannosidase, encoded by the msdS gene, on the cell surface of Y. lipolytica. As the first step to achieve the secretory expression of msdS protein, four different signal sequences-derived from the endogenous Y. lipolytica Lip2 and Xpr2 prepro regions and the heterologous A. niger α-amylase and rice α-amylase signal sequences-were analyzed for their secretion efficiency. It was shown that the YlLip2 prepro sequence was most efficient in directing the secretory expression of msdS in fully N-glycosylated forms. The surface display of msdS was subsequently directed by fusing GPI anchoring motifs derived from Y. lipolytica cell wall proteins, YlCwp1p and YlYwp1p, respectively, to the C-terminus of the Lip2 prepro-msdS protein. The expression of actively functional msdS protein on the cell surface was confirmed by western blot, flow cytometry analysis, along with the α-1,2-mannosidase activity assay using intact Y. lipolytica cells as the enzyme source. Furthermore, the glycoengineered Y. lipolytica Δoch1Δmpo1 strains displaying α-1,2-mannosidase were able to convert Man8GlcNAc2 to Man5GlcNAc2 efficiently on their cell-wall mannoproteins, demonstrating its potential used for glycoengineering in vitro or in vivo.

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Authors and Affiliations

  1. Department of Life Science, Chung-Ang University, Seoul, 156-756, Republic of Korea

    Hye Yun Moon, Trinh Luu Van, Seon Ah Cheon, Jinho Choo & Hyun Ah Kang

  2. Department of Microbiology, School of Bioscience and Biotechnology, Chungnam National University, Daejeon, 305-764, Republic of Korea

    Jeong-Yoon Kim

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  1. Hye Yun Moon
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  2. Trinh Luu Van
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  3. Seon Ah Cheon
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  4. Jinho Choo
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  5. Jeong-Yoon Kim
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  6. Hyun Ah Kang
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Correspondence to Hyun Ah Kang.

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Moon, H.Y., Van, T.L., Cheon, S.A. et al. Cell-surface expression of Aspergillus saitoi-derived functional α-1,2-mannosidase on Yarrowia lipolytica for glycan remodeling. J Microbiol. 51, 506–514 (2013). https://doi.org/10.1007/s12275-013-3344-x

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  • DOI: https://doi.org/10.1007/s12275-013-3344-x

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