Development of a novel rDNA based plasmid for enhanced cell surface display on Yarrowia lipolytica

In this study, a novel rDNA based plasmid was developed for display of heterologous proteins on the cell surface of Yarrowia lipolytica using the C-terminal end of the glycosylphosphatidylinositol (GPI) anchored Y. lipolytica cell wall protein 1 (YlCWP1). mCherry was used as a model protein to asses...

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Bibliographic Details
Published in:AMB Express 2012-05, Vol.2 (1), p.27-27
Main Authors: Bulani, Siyavuya Ishmael, Moleleki, Lucy, Albertyn, Jacobus, Moleleki, Ntsane
Format: Article
Language:English
Subjects:
Biomedical and Life Sciences
Biotechnology
Life Sciences
Microbial Genetics and Genomics
Microbiology
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Summary:In this study, a novel rDNA based plasmid was developed for display of heterologous proteins on the cell surface of Yarrowia lipolytica using the C-terminal end of the glycosylphosphatidylinositol (GPI) anchored Y. lipolytica cell wall protein 1 (YlCWP1). mCherry was used as a model protein to assess the efficiency of the constructed plasmid. Y. lipolytica transformants harbouring the expression cassettes showed a purple colour phenotype on selective YNB-casamino plates as compared to control cells indicating that mCherry was displayed on the cells. Expression of mCherry on cells of Y. lipolytica was confirmed by both fluorescent microscopy and flow cytometry. Furthermore, SDS-PAGE analysis and matrix-assisted laser desorption/ionization (MALDI)-time-of (TOF)-mass spectrometry (MS) peptide mass fingerprinting (PMF) confirmed that the protein cleaved from the yeast cells using enterokinase was mCherry. Efficient cleavage of mCherry reported in this work offers an alternative purification method for displayed heterologous proteins on Y. lipolytica cells using the plasmid constructed in this study. The developed displaying system offers great potential for industrial production and purification of heterologous proteins at low cost.
ISSN:2191-0855
2191-0855
DOI:10.1186/2191-0855-2-27