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A cell surface display system using novel GPI‐anchored proteins in Hansenula polymorpha
Author(s) -
Kim SoYoung,
Sohn JungHoon,
Pyun YuRyang,
Choi EuiSung
Publication year - 2002
Publication title -
yeast
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.923
H-Index - 102
eISSN - 1097-0061
pISSN - 0749-503X
DOI - 10.1002/yea.911
Subject(s) - biology , bacillus subtilis , biochemistry , signal peptide , cell wall , cell , gene , s layer , fusion protein , yeast , microbiology and biotechnology , peptide sequence , bacteria , recombinant dna , genetics
A cell surface display system was developed in yeast Hansenula polymorpha. The four genes HpSED1 , HpGAS1 , HpTIP1 and HpCWP1 , encoding glycosylphosphatidyl‐inositol (GPI)‐anchored cell surface proteins from H. polymorpha , were cloned, characterized and evaluated for their efficacies as cell surface display motifs of reporter proteins. Sequence analysis of these genes revealed that each encodes a typical GPI‐anchored protein that is structurally similar to a counterpart gene in S. cerevisiae . The genes showed a high content of serine‐threonine (alanine) and harboured a putative secretion signal in the N‐terminus and the GPI‐attachment signal in the C‐terminus. The surface anchoring efficiency of these putative cell surface proteins was tested by fusion to the C‐terminal of carboxymethylcellulase (CMCase) from Bacillus subtilis . In all cases, high CMCase activities were detected in intact cell fraction, indicating anchoring of CMCase to the cell surface. HpCwp1p, HpGas1p and the 40 C‐terminal amino acids of HpTip1p from H. polymorpha exhibited a comparatively high CMCase surface anchoring efficiency. When these proteins were used as anchoring motifs for surface display of the glucose oxidase (GOD) from Aspergillus niger , most enzyme activity was detected at the cell surface. Fluorescence activated cell sorter (FACS) analysis of cells displaying GOD on the cell surface demonstrated that GOD was well exposed on the cell surface. HpCwp1p showed the highest anchoring efficiency among others. Copyright © 2002 John Wiley & Sons, Ltd.