High level expression of human MCP-1 using the LCR/MEL expression system.
We have expressed human monocyte chemoattractant protein-1 (hMCP-1) in preerythroid mouse erythroleukemia (MEL) C88 cells using the locus control region/MEL expression system and studied the biological activity of the purified protein in a range of in vitro experimental systems. The recombinant hMCP-1 is expressed at high levels (approximately 10 mg/liter) in this system and is modified in a manner which is very similar to native hMCP-1. We have developed a simple high-yielding two-step purification route employing dye ligand and ion exchange chromatographies which enables us to separate glycosylated and unglycosylated hMCP-1. The purified glycosylated and unglycosylated forms of hMCP-1 have equivalent biological activities in all of the assay systems tested.
Needham
M
M
Vascular, Inflammatory & Musculo-skeletal Research Department, ZENECA Pharmaceuticals, Macclesfield, Cheshire, United Kingdom.
Barratt
D
D
Cerillo
G
G
Green
I
I
Warburton
H
H
Anderson
M
M
Sturgess
N
N
Rollins
B
B
Reilly
C
C
Hollis
M
M
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002118:Calcium / Q000378:metabolism; D018932:Chemokine CCL2 / Q000235:genetics / Q000302:isolation & purification*; D002633:Chemotaxis / Q000235:genetics; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D005786:Gene Expression Regulation / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D006031:Glycosylation; D006801:Humans; D004915:Leukemia, Erythroblastic, Acute / Q000378:metabolism; D013058:Mass Spectrometry; D051379:Mice; D008969:Molecular Sequence Data; D009000:Monocytes / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification; D014162:Transfection / Q000235:genetics; D014407:Tumor Cells, Cultured
D018932:Chemokine CCL2; D017931:DNA Primers; D011994:Recombinant Proteins; D002118:Calcium