Engineering of an FGF-proteoglycan fusion protein with heparin-independent, mitogenic activity

Citation
A. Yoneda et al., Engineering of an FGF-proteoglycan fusion protein with heparin-independent, mitogenic activity, NAT BIOTECH, 18(6), 2000, pp. 641-644
Citations number
15
Categorie Soggetti
Biotecnology & Applied Microbiology",Microbiology
Journal title
NATURE BIOTECHNOLOGY
ISSN journal
10870156 → ACNP
Volume
18
Issue
6
Year of publication
2000
Pages
641 - 644
Database
ISI
SICI code
1087-0156(200006)18:6<641:EOAFFP>2.0.ZU;2-M
Abstract
In the absence of heparan sulfate (HS) on the surface of target cells, or f ree heparin (HP) in the vicinity of their receptors, fibroblast growth fact or (FGF) family members cannot exert their biological activity and are easi ly damaged by proteolysis. This limits the utility of FGFs in a variety of applications including treatment of surgical, burn, and periodontal tissue wounds, gastric ulcers, segmental bony defects, ligament and spinal cord in jury. Here we describe an FGF analog engineered to overcome this limitation by fusing FGF-1 with HS proteoglycan (PG) core protein. The fusion protein (PG-FGF-1), which was expressed in Chinese hamster ovary cells and collect ed from the conditioned medium, possessed both MS and chondroitin sulfate s ugar chains. After fractionation, intact PG-FGF-1 proteins with little affi nity to immobilized HP and high-level HS modification, but not their hepari tinase or heparinase digests, exerted mitogenic activity independent of exo genous MP toward MS-free Ba/F3 transfectants expressing FGF receptor. Altho ugh PG-FGF-1 was resistant to tryptic digestion, its physiological degradat ion with a combination of heparitinase and trypsin augmented its mitogenic activity toward human endothelial cells. The same treatment abolished the a ctivity of simple FGF-1 protein. By constructing a biologically active prot eoglycan-FGF-1 fusion protein, we have demonstrated an approach that may pr ove effective for engineering not only FGF family members, but other MP-bin ding molecules as well.