Development of a novel rDNA based plasmid for enhanced cell surface display on Yarrowia lipolytica
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Authors
Bulani, Siyavuya Ishmael
Moleleki, Lucy Novungayo
Albertyn, Jacobus
Moleleki, Ntsane
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Abstract
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 YNBcasamino
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.
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Keywords
mCherry, rDNA vector, YlCWP1, Yarrowia lipolytica, Cell surface display
Sustainable Development Goals
Citation
Bulani et al.: Development of a novel rDNA based plasmid for enhanced cell surface display on Yarrowia lipolytica. AMB Express 2012 2:27.