High-Level Production of High-Purity Human and Murine Recombinant Prion Proteins Functionally Compatible to In Vitro Seeding Assay
- Authors
- Hwang, Hae-Gwang; Kim, Dae-Hwan; Lee, Jeongmin; Mo, Youngwon; Lee, Se-Hoon; Lee, Yongjin; Hyeon, Jae Wook; Lee, Sol Moe; Cheon, Yong-Pil; Choi, Eun-Kyoung; Kim, Su Yeon; Lee, Yeong Seon; Son, Young-Jin; Ryou, Chongsuk
- Issue Date
- Oct-2018
- Publisher
- 한국미생물·생명공학회
- Keywords
- Expression; high cell-density culture; recombinant prion protein; purification; seeding activity
- Citation
- Journal of Microbiology and Biotechnology, v.28, no.10, pp.1749 - 1759
- Indexed
- SCIE
SCOPUS
KCI
- Journal Title
- Journal of Microbiology and Biotechnology
- Volume
- 28
- Number
- 10
- Start Page
- 1749
- End Page
- 1759
- URI
- https://scholarworks.bwise.kr/erica/handle/2021.sw.erica/5299
- DOI
- 10.4014/jmb.1805.05067
- ISSN
- 1017-7825
- Abstract
- Recombinant (rec) prion protein (PrP) is an extremely useful resource for studying protein misfolding and subsequent protein aggregation events. Here, we report mass production of high-purity rec-polypeptide encoding the C-terminal globular domain of PrP; (90-230) for human and (89-231) for murine PrP. These proteins were expressed as His-tagged fusion proteins in E. coli cultured by a high cell-density aerobic fermentation method. RecPrPs recovered from inclusion bodies were slowly refolded under reducing conditions. Purification was performed by a sequence of metal-affinity, cation-exchange, and reverse-phase chromatography. The current procedure yielded several dozens of milligrams of recPrP per liter with >95% purity. The purified recPrPs predominantly adopted an alpha-helix-rich conformation and were functionally sufficient as substrates to measure the seeding activity of human and animal prions. Establishment of a procedure for high-level production of high-purity recPrP supports the advancement of in vitro investigations of PrP including diagnosis for prion diseases.
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