title: Human granulocyte colony stimulating factor (G-CSF) produced in the filamentous fungus Aspergillus niger.
abstract: For the first time, a fungal production system is described for expression and secretion of the medically important human protein G-CSF, in Aspergillus niger. A reliable strategy was chosen with in-frame fusion of G-CSF behind a KEX2 cleavage site downstream of the coding region of the highly secreted homologous glucoamylase. This provided secretion levels of 5-10 mg/l culture medium of correctly processed G-CSF, although the majority of the protein (>90%) was biologically inactive. Following denaturation/ concentration and chromatographic separation/ renaturation, the G-CSF proliferation activity increased considerably, and analytical immobilised metal affinity chromatography confirmed the monomeric and correctly folded protein. These data suggest that this human secretory protein secreted into the medium of A. niger was not correctly folded, and that it escaped the endoplasmic reticulum folding control systems. This is compared to the folding of G-CSF produced in bacteria and yeast.
authors.forename: Nada
authors.initials: N
authors.lastname: Kraševec
authors.forename: Tatjana
authors.initials: T
authors.lastname: Milunović
authors.forename: Marija Anžur
authors.initials: MA
authors.lastname: Lasnik
authors.forename: Irena
authors.initials: I
authors.lastname: Lukančič
authors.forename: Radovan
authors.initials: R
authors.lastname: Komel
authors.forename: Vladka Gaberc
authors.initials: VG
authors.lastname: Porekar
mesh_terms: D000818:Animals; D001234:Aspergillus niger / Q000378:metabolism*; D001665:Binding Sites; D045744:Cell Line, Tumor; D002845:Chromatography; D002846:Chromatography, Affinity; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D006031:Glycosylation; D016179:Granulocyte Colony-Stimulating Factor / Q000096:biosynthesis*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008670:Metals / Q000737:chemistry; D051379:Mice; D010843:Pichia / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000096:biosynthesis
chemical_list: D008670:Metals; D011994:Recombinant Proteins; D016179:Granulocyte Colony-Stimulating Factor