Switch to
| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
1
|
| pubmed:dateCreated |
1995-6-29
|
| pubmed:abstractText |
Recombinant human interleukin-5 (rhIL-5) was expressed in baculovirus-infected insect cells and purified to homogeneity from the culture medium in a single chromatographic step. Beginning with a cDNA encoding the full-length precursor form of human IL-5, including the authentic secretory leader sequence, recombinant baculovirus-infected insect cells expressed high levels of rhIL-5 (5-15 mg/liter culture) of which > 90% was processed to the mature form and secreted into the culture medium. After removing cells by centrifugation, rhIL-5 was purified by first adjusting the culture medium to the calculated pI value of mature IL-5 (pI 7.44) and then passing the conditioned medium through tandem linked anion- and cation-exchange columns. The resulting pass-through fraction contained the rhIL-5 and was devoid of contaminating proteins. An optional hydrophobic-interaction chromatography step effectively concentrated the pure homodimeric N-glycosylated rhIL-5 with a high overall yield (> 90%). N-terminal amino acid sequence determination indicated that cleavage of the human IL-5 leader sequence in insect cells occurred between Ala19 and Ile20. Recombinant human IL-5 prepared by this procedure bound to the high-affinity IL-5 receptor present on an eosinophilic leukemia cell line and elicited a proliferative response in the IL-5-dependent murine B-cell line BCL1. This rapid and simple procedure for the expression and purification of mature rhIL-5 should therefore enable studies requiring large amounts of this cytokine.
|
| pubmed:language |
eng
|
| pubmed:journal | |
| pubmed:citationSubset |
IM
|
| pubmed:chemical | |
| pubmed:status |
MEDLINE
|
| pubmed:month |
Feb
|
| pubmed:issn |
1046-5928
|
| pubmed:author | |
| pubmed:issnType |
Print
|
| pubmed:volume |
6
|
| pubmed:owner |
NLM
|
| pubmed:authorsComplete |
Y
|
| pubmed:pagination |
63-71
|
| pubmed:dateRevised |
2004-11-17
|
| pubmed:meshHeading |
pubmed-meshheading:7756840-Amino Acid Sequence,
pubmed-meshheading:7756840-Animals,
pubmed-meshheading:7756840-Baculoviridae,
pubmed-meshheading:7756840-Base Sequence,
pubmed-meshheading:7756840-Cell Line,
pubmed-meshheading:7756840-DNA, Complementary,
pubmed-meshheading:7756840-Humans,
pubmed-meshheading:7756840-Interleukin-5,
pubmed-meshheading:7756840-Mice,
pubmed-meshheading:7756840-Molecular Sequence Data,
pubmed-meshheading:7756840-Receptors, Interleukin,
pubmed-meshheading:7756840-Recombinant Proteins,
pubmed-meshheading:7756840-Spodoptera
|
| pubmed:year |
1995
|
| pubmed:articleTitle |
A single-step purification of biologically active recombinant human interleukin-5 from a baculovirus expression system.
|
| pubmed:affiliation |
Department of Biochemistry and Molecular Biology, Merck Frosst Centre for Therapeutic Research, Pointe Claire-Dorval, Quebec, Canada.
|
| pubmed:publicationType |
Journal Article
|