Heparan sulfate oligosaccharides require 6-O-sulfation for promotion of basic fibroblast growth factor mitogenic activity.
The interaction of heparan sulfate (HS) with basic fibroblast growth factor (bFGF) is influential in enabling the growth factor to bind to its cell surface tyrosine kinase receptor. In this study, we have investigated further the structural properties of HS required to mediate the activity of bFGF in a mitogenic assay. We have prepared a library of heparinase III-generated HS oligosaccharides fractionated by both their size (dp6-dp12) and sulfate content. The ability of these oligosaccharides to activate bFGF in a mitogenic assay was then correlated with their length and disaccharide composition. All octa- and hexasaccharide fractions tested were unable to activate bFGF. Dodeca- and decasaccharide fractions were found to contain both activating and non-activating oligosaccharides, and showed a clear correlation between total sulfate content and the level of activatory activity. Disaccharide analysis of a range of dodeca- and decasaccharide fractions showed that both activating and non-activating oligosaccharides were composed mainly of N-sulfated and IdoA(2S)-containing disaccharides. The only significant difference between activating and non-activating oligosaccharides was the content of 6-O-sulfated disaccharides, in particular the disaccharide IdoA(2S)alpha1,4GlcNSO3(6S). These results show that there is a requirement for 6-O-sulfation of N-sulfated glucosamine residues, in addition to the 2-O-sulfation of IdoA, for the promotion of bFGF mitogenic activity by naturally occurring HS oligosaccharides. Analysis of the structure-activity relationships in the dodecasaccharide fractions in particular, suggests that a minimum bFGF activation sequence exists which is dependent on the positioning of at least one 6-O-sulfate group.
CRC Department of Drug Development, Department of Medical Oncology, Paterson Institute for Cancer Research, Christie Hospital, Manchester, M20 4BX United Kingdom. DPlye@picr.man.ac.uk
D A
DA
Pye
R R
RR
Vives
J E
JE
Turnbull
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P
Hyde
J T
JT
Gallagher
D002460:Cell Line; D004187:Disaccharides; D004347:Drug Interactions; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology*; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000187:drug effects; D006497:Heparan Sulfate / Q000737:chemistry / Q000494:pharmacology*; D008221:Lymphoid Tissue / Q000166:cytology / Q000187:drug effects; D008934:Mitogens / Q000494:pharmacology*; D008938:Mitosis / Q000187:drug effects; D009844:Oligosaccharides / Q000737:chemistry / Q000494:pharmacology*; D011133:Polysaccharide-Lyases / Q000494:pharmacology; D013463:Sulfuric Acid Esters / Q000494:pharmacology*
D004187:Disaccharides; D016222:Fibroblast Growth Factor 2; D006497:Heparan Sulfate; D008934:Mitogens; D009844:Oligosaccharides; D011133:Polysaccharide-Lyases; D013463:Sulfuric Acid Esters; C021349:heparitinsulfate lyase