Linked Life Data

9035111

Source:http://linkedlifedata.com/resource/pubmed/id/9035111

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
1
pubmed:dateCreated
1997-4-1
pubmed:abstractText
To create a rapid system to test the effect of sequence changes on recombinant antibody binding, we have developed a procedure for producing functional scFv fragments in an Escherichia coli cell-free translation system. Functional antibodies with antigen-binding activity are obtained only if disulfide formation and rearrangement is allowed to take place during the translation reaction. The inclusion of protein disulfide isomerase (PDI) leads to a threefold increase in yield over that obtained in the presence of glutathione redox systems. DsbA had no such effect, indicating that disulfide shuffing, and not net formation, is the crucial yield-limiting step. The addition of the molecular chaperones DnaK and DnaJ increased the amount of soluble protein but not the amount of functional scFv, which appears to be limited entirely by correct disulfide formation. None of these factors significantly influenced total protein synthesis. In the presence of PDI, chaperones, reduced glutathione and oxidized glutathione, 50% of the scFv produced (about 8 micrograms/ml in only 15 min) could be recovered from immobilized antigen.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jan
pubmed:issn
1087-0156
pubmed:author
pubmed:issnType
Print
pubmed:volume
15
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
79-84
pubmed:dateRevised
2008-11-21
pubmed:meshHeading
pubmed:year
1997
pubmed:articleTitle
Functional antibody production using cell-free translation: effects of protein disulfide isomerase and chaperones.
pubmed:affiliation
Biochemisches Institut, Universität Zürich, Switzerland.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't