Linked Life Data

9226719

Source:http://linkedlifedata.com/resource/pubmed/id/9226719

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
1997-8-18
pubmed:abstractText
The expression of apolipoprotein A-I (apoA-I) has been shown to be very difficult due to its amphiphilic character, autoaggregation, and degradation. We have expressed apoA-I using CHO cells, Baculovirus, and Escherichia coli [Schmidt et al., J. Biol. Chem. (1995) 270, 469-475]. Here we report about optimized conditions for the expression of proapoA-I in CHO cells, testing various serum-free media. We were able to yield apoA-I expression up to 80 micrograms/ml, by far the highest ever reported. However, immunoblot analysis revealed degraded apoA-I. The best apoA-I expression testing various conditions was about 20-30 micrograms/ml without any evidence of degradation. Interestingly, the apoA-I expression resulted in reproducible apoA-I fragments of 26 and 14 kDa. These fragments are consistent with already reported in vivo findings, in which carboxy-terminal proteolysis was suggested. The use of the protease inhibitors pepstatin and chymostatin, both carboxy-peptidase inhibitors, did result in contrast to other studied protease inhibitors in increased apoA-I yield. Therefore, limited carboxy-terminal proteolysis contributes to the degradation of CHO cell-secreted apoA-I. In addition, we evaluated various purification methods for the preparative isolation of recombinant apoA-I. In our hands we obtained the best recovery and no degradation with reversed-phase chromatography using a FPLC system.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jul
pubmed:issn
1046-5928
pubmed:author
pubmed:issnType
Print
pubmed:volume
10
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
226-36
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
1997
pubmed:articleTitle
Expression and purification of recombinant human apolipoprotein A-I in Chinese hamster ovary cells.
pubmed:affiliation
Department of Gastroenterology and Hepatology, Medizinische Hochschule Hannover, Germany.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't