Purification of bioactive glycosylated recombinant glial cell line-derived neurotrophic factor.
Glial cell line-derived neurotrophic factor (GDNF) neuroprotective effect on dopaminergic neurons has been described in vitro and in vivo, turning up as a promising drug for the treatment of Parkinson's disease. Unglycosylated bacteria-obtained GDNF has already been successfully delivered for a long period of time through an infusion pump directly to the putamen of Parkinsonian patients. Nevertheless, improved distribution and safety issues need to be solved and alternative strategies to long-term delivery seem necessary. The use of glycosylated GDNF could eliminate some safety concerns regarding the presence of antibodies against exogenous unglycosylated GDNF used for the treatment. Therefore, we have chosen a mammalian expression system as a source of glycosylated GDNF. In the present work, we describe the purification of recombinant rat GDNF from the culture media of baby hamster kidney (BHK) cells through several purification steps. Highly pure N-glycosylated recombinant GDNF has been obtained similar to the endogenous protein. Furthermore, the purified protein is biologically active when tested its ability to induce PC12 neurite outgrowth.
Garbayo
E
E
Department of Pharmacy and Pharmaceutical Technology, University of Navarra, Irunlarrea, 1, 31080 Pamplona, Spain.
Ansorena
E
E
Lanciego
J L
JL
Aymerich
M S
MS
Blanco-Prieto
M J
MJ
D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D006224:Cricetinae; D051100:Glial Cell Line-Derived Neurotrophic Factor / Q000302:isolation & purification* / Q000494:pharmacology; D006031:Glycosylation; D016501:Neurites / Q000187:drug effects / Q000502:physiology; D051381:Rats; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000494:pharmacology
D051100:Glial Cell Line-Derived Neurotrophic Factor; D011994:Recombinant Proteins