title: A simple and efficient method for the production of human glycosylated glial cell line-derived neurotrophic factor using a Semliki Forest virus expression system.
abstract: Human glial cell line-derived neurotrophic factor (hGDNF) is a very promising protein for the treatment of Parkinson's disease and other neurodegenerative disorders. The present work describes a quick and simple method to obtain a high amount of purified hGDNF using a mammalian cell-derived system. The method is based on the high expression level provided by a Semliki Forest virus vector and its ability to induce a strong shut-off of host-cell protein synthesis in mammalian cells. As a result, hGDNF is the only protein present in the supernatant and can be efficiently purified by a single chromatographic step. Using this system it was possible to eliminate other secreted proteins from the culture medium, like insulin-like growth factor-5, which are hard to remove using other hGDNF production methods. Purified hGDNF presents a complex glycosylation pattern typical of mammalian expression systems and is biologically active. This protocol could be extended to other secreted proteins and could be easily scaled up for industrial purposes.
authors.affiliation: Department of Pharmacy and Pharmaceutical Technology, School of Pharmacy, University of Navarra, C/Irunlarrea 1, 31008 Pamplona, Spain.
authors.forename: Eduardo
authors.initials: E
authors.lastname: Ansorena
authors.forename: Erkuden
authors.initials: E
authors.lastname: Casales
authors.forename: Alejandro
authors.initials: A
authors.lastname: Aranda
authors.forename: Esther
authors.initials: E
authors.lastname: Tamayo
authors.forename: Elisa
authors.initials: E
authors.lastname: Garbayo
authors.forename: Cristian
authors.initials: C
authors.lastname: Smerdou
authors.forename: Maria J
authors.initials: MJ
authors.lastname: Blanco-Prieto
authors.forename: Maria S
authors.initials: MS
authors.lastname: Aymerich
mesh_terms: D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D005347:Fibroblasts; D005822:Genetic Vectors; D051100:Glial Cell Line-Derived Neurotrophic Factor / Q000235:genetics / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D016716:PC12 Cells; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012672:Semliki forest virus / Q000235:genetics*
chemical_list: D051100:Glial Cell Line-Derived Neurotrophic Factor; D011994:Recombinant Proteins