title: Ligation of the fibrin-binding domain by β-strand addition is sufficient for expansion of soluble fibronectin.
abstract: How fibronectin (FN) converts from a compact plasma protein to a fibrillar component of extracellular matrix is not understood. "Functional upstream domain" (FUD), a polypeptide based on F1 adhesin of Streptococcus pyogenes, binds by anti-parallel β-strand addition to discontinuous sets of N-terminal FN type I modules, (2-5)FNI of the fibrin-binding domain and (8-9)FNI of the gelatin-binding domain. Such binding blocks assembly of FN. To learn whether ligation of (2-5)FNI, (8-9)FNI, or the two sets in combination is important for inhibition, we tested "high affinity downstream domain" (HADD), which binds by β-strand addition to the continuous set of FNI modules, (1-5)FNI, comprising the fibrin-binding domain. HADD and FUD were similarly active in blocking fibronectin assembly. Binding of HADD or FUD to soluble plasma FN exposed the epitope to monoclonal antibody mAbIII-10 in the tenth FN type III module ((10)FNIII) and caused expansion of FN as assessed by dynamic light scattering. Soluble N-terminal constructs truncated after (9)FNI or (3)FNIII competed better than soluble FN for binding of FUD or HADD to adsorbed FN, indicating that interactions involving type III modules more C-terminal than (3)FNIII limit β-strand addition to (1-5)FNI within intact soluble FN. Preincubation of FN with mAbIII-10 or heparin modestly increased binding to HADD or FUD. Thus, ligation of FNIII modules involved in binding of integrins and glycosaminoglycans, (10)FNIII and (12-14)FNIII, increases accessibility of (1-5)FNI. Allosteric loss of constraining interactions among (1-5)FNI, (10)FNIII, and (12-14)FNIII likely enables assembly of FN into extracellular fibrils.
authors.affiliation: Department of Biomolecular Chemistry, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA.
authors.forename: Lisa M
authors.initials: LM
authors.lastname: Maurer
authors.forename: Wenjiang
authors.initials: W
authors.lastname: Ma
authors.forename: Nathan L
authors.initials: NL
authors.lastname: Eickstaedt
authors.forename: Ian A
authors.initials: IA
authors.lastname: Johnson
authors.forename: Bianca R
authors.initials: BR
authors.lastname: Tomasini-Johansson
authors.forename: Douglas S
authors.initials: DS
authors.lastname: Annis
authors.forename: Deane F
authors.initials: DF
authors.lastname: Mosher
mesh_terms: D018829:Adhesins, Bacterial / Q000378:metabolism; D000494:Allosteric Regulation / Q000502:physiology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001667:Binding, Competitive / Q000502:physiology; D018604:Epitope Mapping; D005109:Extracellular Matrix / Q000737:chemistry* / Q000378:metabolism*; D005337:Fibrin / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D005353:Fibronectins / Q000737:chemistry* / Q000378:metabolism*; D006493:Heparin / Q000378:metabolism; D006801:Humans; D008024:Ligands; D051379:Mice; D017434:Protein Structure, Tertiary; D012995:Solubility; D013297:Streptococcus pyogenes / Q000378:metabolism
chemical_list: D018829:Adhesins, Bacterial; D000911:Antibodies, Monoclonal; C067625:FN1 protein, human; D005353:Fibronectins; D008024:Ligands; D005337:Fibrin; D006493:Heparin