Recombinant expression of soluble murine prion protein for C-terminal modification

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• 作者：Nam Ky Chu ; Christian F.W. Becker ; ^{christian.becker@univie.ac.at}
• 关键词：ASP ; 6xHis-ATPase-SrtA-PrP ; ASPM ; 6xHis-ATPase-SrtA-PrP-MxeIntein-6xHis-CBD ; CBD ; chitin binding domain ; GPI ; glycosylphosphatidylinositol ; GSH ; reduced glutathione ; GSSG ; oxidized glutathione ; HDP ; 6xHis-DnaK-TEV-PrP ; IPTG ; isopropyl-¦Â-d-thiogalactopyra
• 刊名：FEBS Letters
• 出版年：2013
• 出版时间：1 March, 2013
• 年：2013
• 卷：587
• 期：5
• 页码：430-435
• 全文大小：772 K

文摘

Membrane attachment of prion protein (PrP) via its glycosylphosphatidylinositol (GPI) anchor plays a key role during conversion of cellular PrP^C into its pathogenic isoform PrP^Sc. Strategies to access homogenous lipidated PrP via expressed protein ligation (EPL) are required to fully decipher the effect of membrane attachment. Such strategies suffer from insoluble expression of PrP-intein fusion constructs and low folding efficiencies that severely limit the available amount of homogeneous lipidated PrP. Here, we describe an alternative method for expression of soluble PrP-intein fusion proteins in Escherichia coli that provides access to natively folded PrP ready to use in EPL.