Characterization of growth factor-binding structures in heparin/heparan sulfate using an octasaccharide library.
Heparan sulfate (HS) chains interact with various growth and differentiation factors and morphogens, and the most interactions occur on the specific regions of the chains with certain monosaccharide sequences and sulfation patterns. Here we generated a library of octasaccharides by semienzymatic methods by using recombinant HS 2-O-sulfotransferase and HS 6-O-sulfotransferase, and we have made a systematic investigation of the specific binding structures for various heparin-binding growth factors. An octasaccharide (Octa-I, DeltaHexA-GlcNSO(3)-(HexA-GlcNSO(3))(3)) was prepared by partial heparitinase digestion from completely desulfated N-resulfated heparin. 2-O- and 6-O-sulfated Octa-I were prepared by enzymatically transferring one to three 2-O-sulfate groups and one to three 6-O-sulfate groups per molecule, respectively, to Octa-I. Another octasaccharide containing 3 units of HexA(2SO(4))-GlcNSO(3)(6SO(4)) was prepared also from heparin. This octasaccharide library was subjected to affinity chromatography for interactions with fibroblast growth factor (FGF)-2, -4, -7, -8, -10, and -18, hepatocyte growth factor, bone morphogenetic protein 6, and vascular endothelial growth factor, respectively. Based upon differences in the affinity to those octasaccharides, the growth factors could be classified roughly into five groups: group 1 needed 2-O-sulfate but not 6-O-sulfate (FGF-2); group 2 needed 6-O-sulfate but not 2-O-sulfate (FGF-10); group 3 had the affinity to both 2-O-sulfate and 6-O-sulfate but preferred 2-O-sulfate (FGF-18, hepatocyte growth factor); group 4 required both 2-O-sulfate and 6-O-sulfate (FGF-4, FGF-7); and group 5 hardly bound to any octasaccharides (FGF-8, bone morphogenetic protein 6, and vascular endothelial growth factor). The approach using the oligosaccharide library may be useful to define specific structures required for binding to various heparin-binding proteins. Octasaccharides with the high affinity to FGF-2 and FGF-10 had the activity to release them, respectively, from their complexes with HS. Thus, the library may provide new reagents to specifically regulate bindings of the growth factors to HS.
Ashikari-Hada
Satoko
S
Institute for Molecular Science of Medicine, Aichi Medical University, Nagakute, Aichi 480-1195, Japan.
Habuchi
Hiroko
H
Kariya
Yutaka
Y
Itoh
Nobuyuki
N
Reddi
A Hari
AH
Kimata
Koji
K
D000595:Amino Acid Sequence; D055418:Bone Morphogenetic Protein 6; D019485:Bone Morphogenetic Proteins / Q000378:metabolism; D002454:Cell Differentiation; D002846:Chromatography, Affinity; D004187:Disaccharides / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D051526:Fibroblast Growth Factor 10; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism; D005346:Fibroblast Growth Factors / Q000378:metabolism; D006133:Growth Substances / Q000737:chemistry* / Q000378:metabolism; D006493:Heparin / Q000737:chemistry*; D006497:Heparan Sulfate / Q000737:chemistry*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D011134:Polysaccharides / Q000737:chemistry*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D012685:Sepharose / Q000494:pharmacology; D017386:Sequence Homology, Amino Acid; D012965:Sodium Chloride / Q000494:pharmacology; D020349:Surface Plasmon Resonance
D055418:Bone Morphogenetic Protein 6; D019485:Bone Morphogenetic Proteins; D004187:Disaccharides; D051526:Fibroblast Growth Factor 10; D016222:Fibroblast Growth Factor 2; D005346:Fibroblast Growth Factors; D006133:Growth Substances; D006493:Heparin; D006497:Heparan Sulfate; D011134:Polysaccharides; D011994:Recombinant Proteins; D012685:Sepharose; D012965:Sodium Chloride; C525816:BMP6 protein, human; C496463:FGF10 protein, human