A. Yoneda et al., Engineering of an FGF-proteoglycan fusion protein with heparin-independent, mitogenic activity, NAT BIOTECH, 18(6), 2000, pp. 641-644
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.