Linked Life Data

17523923

Source:http://linkedlifedata.com/resource/pubmed/id/17523923

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
Pt 4
pubmed:dateCreated
2007-11-13
pubmed:abstractText
Dilution refolding of recombinant consensus IFN (interferon) from inclusion bodies suffers from low yield. A stable intermediate was found to mix with the correct product and to have an antiviral activity of less than 10% of the latter. This intermediate would form precipitates upon removal of the precipitation inhibitor arginine. Compared with the native protein, the intermediate moved more slowly on non-reducing SDS/PAGE. The CD and fluorescence spectra indicated that it had formed a native-like structure, but had only one disulfide bond: Cys(29)-Cys(139). Further evidence showed that the formation of Cys(29)-Cys(139) is specific and very likely to happen, even in the presence of a high concentration of reducing agent, whereas pairing of the other disulfide (Cys(1)-Cys(99)) needed a stronger oxidative condition. It competed with intermolecular disulfide bonding to form covalent oligomers. On the basis of this discovery, a two-stage refolding step strategy was designed that employed a modified dilution refolding step followed by a dialysis refolding step. The first stage used a high concentration of reducing agent together with the precipitation inhibitor arginine. The purpose was to hinder any reaction through Cys(1) or Cys(99) but allow the intramolecular disulfide bonding of Cys(29)-Cys(139). The second stage was a dialysis step that gradually increased the oxidative agent concentration and simultaneously decreased the arginine concentration. The refolding yield was increased from 35 to 82%, while the mass recovery was increased from 60 to 96%. Moreover, this strategy could suppress precipitation even after arginine was completely removed.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Dec
pubmed:issn
1470-8744
pubmed:author
pubmed:issnType
Electronic
pubmed:volume
48
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
189-98
pubmed:dateRevised
2011-11-17
pubmed:meshHeading
pubmed:year
2007
pubmed:articleTitle
Identification of an oxidative refolding intermediate of recombinant consensus interferon from inclusion bodies and design of a two-stage strategy to promote correct disulfide-bond formation.
pubmed:affiliation
National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beiertiao 1, Zhongguancun, Beijing 100080, People's Republic of China.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't