Yeast arming by the Aga2p system: effect of growth conditions in galactose on the efficiency of the display and influence of expressing leucine-containing peptides
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The amino or carboxy-terminal regions of certain cell wall proteins are capable of anchoring foreign proteins or peptides on the cell wall of the yeast Saccharomyces cerevisiae. This possibility has resulted in the development of a methodology known as yeast display which has powerful applications in biotechnology, pharmacy, and medicine. This work describes the results of experiments in which the agglutinin Aga2p protein is used as an anchor and several leucine-based peptides have been introduced into its N-terminal or C-terminal position. We found that the sequence of these peptides can affect plasmid stability, growth kinetics, and levels of the fusion protein displayed, and we analyzed how the incubation conditions influence these parameters. Besides, we show that the introduction of these small peptides can modify the properties of cell cover; in particular, fusing five or ten leucine residues to the Aga2p protein results in greater hydrophobicity of the cell wall and also in increased resistance to the presence of the organic solvents acetonitrile and ethanol and to high salt concentrations. The introduction of the RLRLL sequence also results in higher resistance to the exposure of yeast cells to NaCl stress.
KeywordsAga2 protein Cell wall hydrophobicity Short peptides Stress resistance Yeast display
We are indebted to Drs. Wittrup and Neville for providing us with the pCTCON2 and pYD5 plasmids, respectively. We gratefully acknowledge SCSIE (Universitat de València) for access to its instrumental facilities of DNA sequencing, flow cytometry, and electron microscopy. This work has been supported by grants from the Spanish Ministry of Science and Technology BFU2008-04082-C02-01/BMC and BFU2011-23501/BMC.
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