STRUCTURE/FUNCTION OF BONE GLYCOPROTEINS

骨是由多种细胞类型、细胞外基质和 矿物质正常的功能取决于协调的努力， 几种细胞类型，但它是成骨细胞的影响下，当地 和系统因素，主要负责合成， 细胞外基质的组织和沉积， 在骨形成过程中矿化。当再吸收和 骨基质的沉积被破坏，各种代谢骨 诸如骨质疏松症、佩吉特病和骨硬化症之类的疾病可以 结果.这些研究的目的是确定 骨形成中的细胞外基质沉积。这些研究将导致 更好地理解骨稳态，从而具有广泛的 代谢性骨疾病谱和病理学的应用 结缔组织。 该项目的具体目标是应用尖端分子和 细胞生物学技术来确定双糖蛋白聚糖和骨的作用 唾液酸蛋白在骨细胞外基质形成中的作用。许多 骨基质的蛋白质经历了大量的翻译后 修饰包括糖基化、磷酸化和硫酸化。 生物测定受到变性要求的阻碍， 纯化过程中的溶剂和组织的固有产率差。到 生产后处理的重组糖蛋白， 化学量，我们将开发一种新的系统， 在成骨样细胞中表达。这项研究的重要进展 将使复杂的翻译后的作用， 这些分子的修饰被定义。这个项目的最终目标 研究的目的是确定这些分子的功能， 与细胞外基质、驻留细胞和生长的相互作用 因素这将通过（a）短暂的、短期的表达和 (b)野生型和突变型细胞外 基质糖蛋白分子间相互作用将通过以下方法进行研究（a） 成骨细胞培养物中存款成熟的细胞内表达 胶原性细胞外基质和（B）通过体外结合研究， 主要基于可在本发明中获得的表面等离子体共振技术， BIAcore系统（Pharmacia）。

Bone is a composite of diverse cell types, extracellular matrix and mineral. Normal function is dependent upon the co-ordinate effort of several cell types, but it is the osteoblast under the influence of local and systemic factors that is primarily responsible for the synthesis, organization and deposition of an extracellular matrix that becomes mineralized during bone formation. When the balance between resorption and deposition of the bone matrix is disrupted, a variety of metabolic bone diseases such as osteoporosis, Paget's disease, and osteopetrosis can result. The goal of these studies is to define the role of the extracellular matrix deposition in bone formation. These studies will lead to a greater understanding of bone homeostasis and thereby have broad application to the spectrum of metabolic bone diseases and the pathology of connective tissues. The specific aim of this project is to apply cutting edge molecular and cell biology techniques to define the role of biglycan and bone sialoprotein in the formation of the bone extracellular matrix. Many of the proteins of the bone matrix undergo extensive post-translational modifications including glycosylation, phosphorylation and sulfation. Biological assays have been hampered by the requirement for denaturing solvents during purification and inherently poor yields from tissues. To produce post-translationally processed recombinant glycoproteins in chemical amounts, we will develop a novel system for vaccinia virus-driven expression in osteoblast-like cells. This important advance in the study of matrix biology will enable the role of the complex post-translational modifications of these molecules to be defined. The ultimate goal of this research is to determine the function of these molecules and define their interaction with the extracellular matrix, the resident cells, and growth factors. This will be achieved by (a) transient, short-term expression and (b) stable, long-term expression of wild-type and mutant extracellular matrix glycoproteins. Intermolecular interactions will be studied (a) by in cellulo expression in osteoblast cultures that deposit mature collagenous extracellular matrices and (b)by in vitro binding studies, primarily based on the technique of surface plasmon resonance available in the BIAcore System (Pharmacia).