title: Bone acidic glycoprotein-75 self-associates to form macromolecular complexes in vitro and in vivo with the potential to sequester phosphate ions.
abstract: Monoclonal antibody HTP IV-#1 specifically recognizes a complexation-dependent neoepitope on bone acidic glycoprotein-75 (BAG-75) and a Mr = 50 kDa fragment. Complexes of BAG-75 exist in situ, as shown by immunofluorescent staining of the primary spongiosa of rat tibial metaphysis and osteosarcoma cell micromass cultures with monoclonal antibody HTP IV-#1. Incorporation of BAG-75 into complexes by newborn growth plate and calvarial tissues was confirmed with a second, anti-BAG-75 peptide antibody (#503). Newly synthesized BAG-75 immunoprecipitated from mineralizing explant cultures of bone was present entirely in large macromolecular complexes, while immunoprecipitates from monolayer cultures of osteoblastic cells were previously shown to contain only monomeric Mr = 75 kDa BAG-75 and a 50 kDa fragment. Purified BAG-75 self-associated in vitro to form large spherical aggregate structures composed of a meshwork of 10 nm diameter fibrils. These structures have the capacity to sequester large amounts of phosphate ions as evidenced by X-ray microanalysis and by the fact that purified BAG-75 preparations, even after extensive dialysis against water, retained phosphate ions in concentrations more than 1,000-fold higher than can be accounted for by exchange calculations or by electrostatic binding. The ultrastructural distribution of immunogold-labeled BAG-75 in the primary spongiosa underlying the rat growth plate is distinct from that for other acidic phosphoproteins, osteopontin and bone sialoprotein. We conclude that BAG-75 self-associates in vitro and in vivo into microfibrillar complexes which are specifically recognized by monoclonal antibody HTP IV-#1. This propensity to self-associate into macromolecular complexes is not shared with acidic phosphoproteins osteopontin and bone sialoprotein. We hypothesize that an extracellular electronegative network of macromolecular BAG-75 complexes could serve an organizational role in forming bone or as a barrier restricting local diffusion of phosphate ions.
authors.affiliation: Division of Molecular Biology and Biochemistry, School of Biological Sciences, University of Missouri-KC, Kansas City 64110, USA.
authors.forename: J P
authors.initials: JP
authors.lastname: Gorski
authors.forename: E A
authors.initials: EA
authors.lastname: Kremer
authors.forename: Y
authors.initials: Y
authors.lastname: Chen
authors.forename: S
authors.initials: S
authors.lastname: Ryan
authors.forename: C
authors.initials: C
authors.lastname: Fullenkamp
authors.forename: J
authors.initials: J
authors.lastname: Delviscio
authors.forename: K
authors.initials: K
authors.lastname: Jensen
authors.forename: M D
authors.initials: MD
authors.lastname: McKee
mesh_terms: D000818:Animals; D001842:Bone and Bones / Q000737:chemistry* / Q000276:immunology / Q000648:ultrastructure; D003460:Crystallization; D019281:Dimerization; D000939:Epitopes / Q000737:chemistry; D006023:Glycoproteins / Q000737:chemistry* / Q000276:immunology; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D010710:Phosphates / Q000737:chemistry*; D011485:Protein Binding; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D000939:Epitopes; D006023:Glycoproteins; D046911:Macromolecular Substances; D010710:Phosphates; C512787:bone acidic glycoprotein-75, rat