Purification and molecular cloning of prostacyclin-stimulating factor from serum-free conditioned medium of human diploid fibroblast cells.
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Yamauchi T, Umeda F, Masakado M, Isaji M, Mizushima S, Nawata H
Purification and molecular cloning of prostacyclin-stimulating factor from serum-free conditioned medium of human diploid fibroblast cells.
Biochem J. 1994 Oct 15;303 ( Pt 2):591-8.
- PubMed ID
- 7980422 [ View in PubMed]
- Abstract
We attempted to identify the factor that stimulated prostacyclin (PGI2) production using conditioned medium from cultured human diploid fibroblast cells subjected to a series of purification steps using h.p.l.c. on DEAE-5PW, Heparin-5PW, Protein-Pak 300, and an insulin-like growth factor-1 ligand affinity column. The purified prostacyclin-stimulating factor (PSF) ran as a single band with a molecular mass of 31 kDa by SDS/PAGE. Analysis of the purified PSF by C4 reversed-phase h.p.l.c. showed a single sharp peak in 31% (v/v) acetonitrile. The material was purified 8000-fold with an overall yield of about 18%. The purified PSF stimulated PGI2 production by cultured bovine aortic endothelial cells at a concentration of about 10 ng/ml; maximal stimulation was achieved at a concentration of 25 ng/ml. A cDNA coding for PSF was cloned and sequenced, revealing an apparently novel protein with no obvious sequence similarity to known proteins.