title: Characterization of putative glycosylphosphatidylinositol-anchoring motifs for surface display in the methylotrophic yeast Hansenula polymorpha.
abstract: Bioinformatic analysis of the genome of the methylotrophic yeast Hansenula polymorpha revealed 39 putative glycosylphosphatidylinositol-anchored proteins (GPI-proteins). Notably, dibasic motifs in the proximal ω-site, that has been reported as a plasma membrane retention signal in Saccharomyces cerevisiae GPI-proteins, were not found in any of the predicted GPI-proteins of H. polymorpha. To evaluate the in silico prediction, C-terminal peptides of 40 amino acids derived from ten H. polymorpha GPI-proteins were fused to the Aspergillus saitoi α-1,2-mannosidase (msdS). Cell wall fraction analysis showed that nine of the ten msdS-GPI fusion proteins were mostly localized at the cell wall. Surface expression of functional msdS was further confirmed by in vitro enzyme activity assay and by glycan structure analysis of cell wall mannoproteins. The recombinant H. polymorpha strains expressing surface-displayed msdS have the potential as useful hosts to produce glycoproteins with decreased mannosylation.
authors.affiliation: Department of Life Science, College of Natural Science, Chung-Ang University, Seoul, 156-756, Korea.
authors.forename: Seon Ah
authors.initials: SA
authors.lastname: Cheon
authors.forename: Jinhee
authors.initials: J
authors.lastname: Jung
authors.forename: Jin Ho
authors.initials: JH
authors.lastname: Choo
authors.forename: Doo-Byoung
authors.initials: DB
authors.lastname: Oh
authors.forename: Hyun Ah
authors.initials: HA
authors.lastname: Kang
mesh_terms: D001665:Binding Sites; D002473:Cell Wall / Q000378:metabolism*; D003001:Cloning, Molecular; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D010843:Pichia / Q000737:chemistry / Q000166:cytology / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D043323:alpha-Mannosidase / Q000378:metabolism
chemical_list: D005656:Fungal Proteins; D017261:Glycosylphosphatidylinositols; D008562:Membrane Glycoproteins; D011993:Recombinant Fusion Proteins; D043323:alpha-Mannosidase