Dentin Matrix Protein 1 Immobilized on Type I Collagen Fibrils Facilitates Apatite Deposition in Vitro

Abstract

During bone and dentin mineralization, the crystal nucleation and growth processes are considered to be matrix regulated. Osteoblasts and odontoblasts synthesize a polymeric collagenous matrix, which forms a template for apatite initiation and elongation. Coordinated and controlled reaction between type I collagen and bone/dentin-specific noncollagenous proteins are necessary for well defined biogenic crystal formation. However, the process by which collagen surfaces become mineralized is not understood. Dentin matrix protein 1 (DMP1) is an acidic noncollagenous protein expressed during the initial stages of mineralized matrix formation in bone and dentin. Here we show that DMP1 bound specifically to type I collagen, with the binding region located at the N-telopeptide region of type I collagen. Peptide mapping identified two acidic clusters in DMP1 responsible for interacting with type I collagen. The collagen binding property of these domains was further confirmed by site-directed mutagenesis. Transmission electron microscopy analyses have localized DMP1 in the gap region of the collagen fibrils. Fibrillogenesis assays further demonstrated that DMP1 accelerated the assembly of the collagen fibrils in vitro and also increased the diameter of the reconstituted collagen fibrils. In vitro mineralization studies in the presence of calcium and phosphate ions demonstrated apatite deposition only at the collagenbound DMP1 sites. Thus specific binding of DMP1 and possibly other noncollagenous proteins on the collagen fibril might be a key step in collagen matrix organization and mineralization.Mineralized tissues such as bone and dentin are hierarchically organized biocomposites possessing unique mechanical properties (1). Understanding the biomineralization process is of great value for synthesis of bioengineered bone and dentinlike materials with optimum structural properties. Type I collagen accounts for 90% of the total protein in the organic matrix of bone and dentin (1). It not only provides the structural framework with viscoelastic properties but also defines compartments for ordered mineral deposition. Studies have demonstrated that the apatite crystals are first nucleated in the gap region, and the growing mineral platelets are highly organized in a staggered manner within the collagen fibrils (2, 3).It is well established that type I collagen matrix does not have the capacity to induce matrix-specific mineral formation from metastable calcium phosphate solutions that do not spontaneously precipitate. Several in vitro studies have demonstrated that ordered mineralization of apatite on collagen fibril is impossible without additives (4, 5). As a result, much attention has been drawn to the noncollagenous proteins (NCPs) 1 that are tightly bound to the collagen fibers in mineralized tissues. Bone and/or dentin-specific NCPs are mostly acidic in nature and are rich in glutamic acid, aspartic acid, and phosphoserines (6, 7). They possess high calcium binding capacity and hydroxyapatite affinity (8 -10). In vitro ...

Stress Chaperone GRP-78 Functions in Mineralized Matrix Formation

“…Recently, we have demonstrated that dentin matrix protein 1 (DMP1), an acidic phosphoprotein, initially identified from the extracellular dentin matrix and implicated in differentiation of mesenchymal cells and also in biomineralization (13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23), binds to GRP-78 found on the plasma membrane of odontoblasts and is endocytosed via the caveolar pathway (24). In the present study we show that a similar physiological process occurs in pre-osteoblasts and osteoblasts.…”

“…The rationale was to mimic the ECM environment when both DMP1 and GRP-78 are available to bind to type I collagen and to each other. The ability of DMP1 to bind to type I collagen and aid nucleation of hydroxyapatite has been shown previously (19). Results showed a two-phase binding wherein GRP-78 bound type I collagen first, as it had a higher binding affinity (comparing data from Figs.…”

Immunohistochemical localization of dentin matrix protein 1 in human dentin

“…DMP1 has been proposed to possess dual functions, both as a transcription factor that targeted the nucleus and as an extracellular matrix protein that initiated mineralization (Narayanan et al 2003;. Cooperative interactions between DMP1 and type I collagen can initiate apatite nucleation and mineral deposition (He & George 2004, He et al 2005. Recently, in vivo studies by Lu and coworkers (Lu et al 2007) revealed that DMP1-null mice show abnormalities in mineralization and structural organization of dentin.…”

The enigmas of bone without osteocytes

“…[7][8][9][10] The extensiveness of the lacunar-canalicular network provides support for the hypothesis that osteocytes function primarily as local sensors, triggering processes that allow bone to respond to surrounding environmental mechanical conditions or to hormonal signals during changing ion homeostasis demands. As the mechanosensors of bone, osteocytes detect local changes in strain in their vicinity, [11][12][13] and in response express membranebound proteins and release soluble factors that regulate and coordinate the function of bone surface cells (bone-forming osteoblasts and bone-resorbing osteoclasts 3,[14][15][16][17] ). The critical role of osteocytes in mechanosensing and bone tissue health is underlined for instance by Tatsumi et al's study, 18 where targeted ablation of osteocytes in a transgenic mouse model induced osteoporosis and a reduced response to changing mechanoenvironment.…”