Single-step purification and refolding of recombinant mouse and human myelin oligodendrocyte glycoprotein and induction of EAE in mice.
The extracellular domain of human and rat MOG (ED-MOG) induces experimental autoimmune encephalomyelitis (EAE) when injected into susceptible animals. EAE is a T cell-mediated disease of the central nervous system commonly used as an animal model for human multiple sclerosis. Here, we describe a straightforward procedure for the purification and refolding of mouse and human ED-MOG overexpressed in Escherichia coli as inclusion bodies. Following solubilization and purification using Ni-NTA resin chromatography under denaturing conditions, a column-based refolding proceeded in renaturation buffer supplemented with a glutathione redox buffer system. Using this approach up to 33 mg of highly pure soluble proteins was obtained per liter of expression culture. The ability of purified proteins to induce EAE was evaluated in three strains of mice. We believe that the strategy described here would facilitate researchers to carry out encephalitogenic as well as structure-function studies of this autoantigen. Additionally, we show for the first time that mouse ED-MOG induces severe disease in mice.
Liñares
David
D
Dupont Performance Coatings Iberica, S.L. Barcelona 08213, Spain.
Echevarria
Iñigo
I
Mañá
Paula
P
D000818:Animals; D002845:Chromatography; D004681:Encephalomyelitis, Autoimmune, Experimental / Q000139:chemically induced / Q000276:immunology*; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics; D006801:Humans; D002479:Inclusion Bodies / Q000737:chemistry; D051379:Mice; D008810:Mice, Inbred C57BL; D009185:Myelin Proteins; D018992:Myelin-Associated Glycoprotein / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D063308:Myelin-Oligodendrocyte Glycoprotein; D017510:Protein Folding; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification
C571326:MOG protein, human; C571327:Mog protein, mouse; C571328:Mog protein, rat; D009185:Myelin Proteins; D018992:Myelin-Associated Glycoprotein; D063308:Myelin-Oligodendrocyte Glycoprotein; D011994:Recombinant Proteins