title: Towards a resolution of the stoichiometry of the fibroblast growth factor (FGF)-FGF receptor-heparin complex.
abstract: The 22 members of the fibroblast growth factor (FGF) family have been implicated in cell proliferation, differentiation, survival, and migration. They are required for both development and maintenance of vertebrates, demonstrating an exquisite pattern of affinities for both protein and proteoglycan receptors. Recent crystal structures have suggested two models for the complex between FGFs, FGF receptors (FGFRs) and the proteoglycan heparan sulphate that mediates signalling, and have provided insight into how FGFs show differing affinities for the range of FGFRs. However, the physiological relevance of the two different models has not been made clear. Here, we demonstrate that the two complexes can be prepared from the same protein components, confirming that neither complex is the product of misfolded protein samples. Analyses of the complexes with mass spectrometry and analytical ultracentrifugation show that the species observed are consistent with the crystal structures formed using the two preparation protocols. This analysis supports the contention that both of the crystal structures reflect the state of the molecules in solution. Mass spectrometry of the complexes suggests that the stoichiometry of the complexes is 2 FGF1:2 FGFR2:1 heparin, regardless of the method used to prepare the complexes. These observations suggest that the two proposed complex architectures may both have relevance to the formation of an in vivo signalling complex, with a combination of the two interactions contributing to the formation of a larger focal complex.
authors.affiliation: Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, UK. nic@cryst.bioc.cam.ac.uk
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Harmer
authors.forename: Leopold L
authors.initials: LL
authors.lastname: Ilag
authors.forename: Barbara
authors.initials: B
authors.lastname: Mulloy
authors.forename: Luca
authors.initials: L
authors.lastname: Pellegrini
authors.forename: Carol V
authors.initials: CV
authors.lastname: Robinson
authors.forename: Tom L
authors.initials: TL
authors.lastname: Blundell
mesh_terms: D002850:Chromatography, Gel; D005346:Fibroblast Growth Factors / Q000737:chemistry / Q000378:metabolism*; D006493:Heparin / Q000737:chemistry / Q000378:metabolism*; D013058:Mass Spectrometry; D008958:Models, Molecular; D017468:Receptors, Fibroblast Growth Factor / Q000737:chemistry / Q000378:metabolism*; D014461:Ultracentrifugation
chemical_list: D017468:Receptors, Fibroblast Growth Factor; D005346:Fibroblast Growth Factors; D006493:Heparin