title: Construction of a novel synergistic system for production and recovery of secreted recombinant proteins by the cell surface engineering.
abstract: We determined whether the cocultivation of yeast cells displaying a ZZ-domain and secreting an Fc fusion protein can be a novel tool for the recovery of secreted recombinant proteins. The ZZ-domain from Staphylococcus aureus protein A was displayed on the cell surface of Saccharomyces cerevisiae under the control of the GAL1 promoter. Strain S. cerevisiae BY4742 cells displaying the ZZ-domain on their surface were used for cocultivation with cells that produce a target protein fused to the Fc fragment as an affinity tag. The enhanced green fluorescent protein or Rhizopus oryzae lipase was genetically fused to the N and C termini of the Fc fragment of human immunoglobulin G, respectively. Through analysis by fluorescence-activated cell sorting and enzymatic assay, it was demonstrated that these fusion proteins are successfully produced in the medium and recovered by affinity binding with the cell surface displaying the ZZ-domain. These results suggest that the ZZ-domain-displaying cell and Fc fusion protein-secreting cell can be applied to use in synergistic process of production and recovery of secreted recombinant proteins.
authors.affiliation: Department of Applied Chemistry, Kobe City College of Technology, Gakuenhigashi, Nishi-ku, Kobe, 651-2194, Japan. gfp@kobe-kosen.ac.jp
authors.forename: Seiji
authors.initials: S
authors.lastname: Shibasaki
authors.forename: Ai
authors.initials: A
authors.lastname: Kawabata
authors.forename: Jun
authors.initials: J
authors.lastname: Ishii
authors.forename: Shunsuke
authors.initials: S
authors.lastname: Yagi
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Kadonosono
authors.forename: Michiko
authors.initials: M
authors.lastname: Kato
authors.forename: Nobuo
authors.initials: N
authors.lastname: Fukuda
authors.forename: Akihiko
authors.initials: A
authors.lastname: Kondo
authors.forename: Mitsuyoshi
authors.initials: M
authors.lastname: Ueda
mesh_terms: D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D015870:Gene Expression; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008049:Lipase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D015202:Protein Engineering / Q000379:methods*; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012233:Rhizopus / Q000201:enzymology; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D013205:Staphylococcal Protein A / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D005656:Fungal Proteins; D007141:Immunoglobulin Fc Fragments; D011993:Recombinant Fusion Proteins; D013205:Staphylococcal Protein A; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D008049:Lipase