STRUCTURE/FUNCTION OF BONE GLYCOPROTEINS

骨是由不同类型的细胞、细胞外基质和 矿物。正常的功能依赖于 几种细胞类型，但它是成骨细胞受局部影响 以及主要负责合成的系统性因素， 细胞外基质的组织和沉积 在骨形成过程中矿化。当再吸收和再吸收之间的平衡 骨基质的沉积被打乱，各种新陈代谢的骨 骨质疏松症、佩吉特氏病和骨化症等疾病可能 结果。这些研究的目标是确定 骨形成中的细胞外基质沉积。这些研究将导致 对骨骼动态平衡有了更好的了解，从而有了广泛的 代谢性骨病谱与病理学的应用 结缔组织。 该项目的具体目标是应用尖端的分子和 确定二聚糖和骨的作用的细胞生物学技术 唾液酸蛋白参与骨细胞外基质的形成。许多. 骨基质的蛋白质经历了广泛的翻译后 修饰包括糖基化、磷酸化和硫酸盐化。 生物检测由于变性的要求而受到阻碍。 在提纯过程中使用溶剂，从组织中提取的产率本来就很低。至 产生翻译后加工的重组糖蛋白 化学量，我们将开发一种新型的牛痘病毒驱动系统 在成骨细胞样细胞中表达。这项研究的重要进展 基质生物学的作用将使复杂的翻译后 这些分子的修饰有待定义。这样做的最终目的是 研究是确定这些分子的功能并定义它们的 与细胞外基质、驻留细胞和生长的相互作用 各种因素。这将通过(A)瞬时的、短期的表达和 (B)野生型和突变型胞外的稳定、长期表达 基质糖蛋白。分子间的相互作用将研究(A) 在沉积成熟的成骨细胞培养中的细胞表达 胶原细胞外基质和(B)通过体外结合研究， 主要基于可用在 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).