Expression, purification and characterisation of secreted recombinant glycoprotein PsA in Dictyostelium discoideum

Ti Zhou-Chou, Martin B. Slade, Keith L. Williams*, Andrew A. Gooley

*Corresponding author for this work

Research output: Contribution to journalArticle

9 Citations (Scopus)


Dictyostelium discoideum is a newly developed eukaryotic expression system which is an alternative to tissue cultures for the production of recombinant proteins requiring eukaryotic folding and post-translational modifications. The homologous glycoprotein PsA (prespore specific antigen) is a glycosyl phosphatidylinositol (GPI) anchored membrane protein from D. discoideum. A truncated form of PsA has been expressed in D. discoideum and secreted into a peptone based broth at levels of 10 mg per l growth medium. A simple purification protocol for recombinant PsA (rPsA) involved three steps: the concentration of the culture supernatant by ammonium sulfate precipitation, Mono Q anion-exchange chromatography, followed by size exclusion chromatography on Superdex 75. 20 mg of rPsA was purified to 98% purity from 3.7 1 culture supernatant. Purified rPsA was characterised. The molecular mass of the purified rPsA is 15.6 kDa, which suggests that the molecule is secreted as a monomer and contains 12% (w/w) carbohydrate. The protein sequence of rPsA proved identical to that of the predicted DNA construct. Although the recombinant form of PsA is expressed at a different developmental stage from the native molecule, the same Thr residues that are O-glycosylated in the authentic molecule are glycosylated in the recombinant protein.

Original languageEnglish
Pages (from-to)137-149
Number of pages13
JournalJournal of Biotechnology
Issue number2
Publication statusPublished - 15 Jan 1995


  • Dictyostelium discoideum
  • Eukaryote expression system
  • Recombinant glycoprotein
  • Recombinant protein purification

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