PURIFICATION OF A BONE NUCLEAR MATRIX PROTEIN

描述：组织基质是连接骨骼的动态前沿 细胞核和细胞质与细胞外基质的网络。 这些研究旨在检验核基质的假设 蛋白4(NMP4)是一种新的甲状旁腺素反应的成骨细胞核基质因子， 在促进1型胶原α1链的表达中起作用 (COLA1)在成骨细胞中。这些研究的主要目的是净化 NMP4，用于获得生成所需的微测序数据 这种新蛋白的抗体。核基质蛋白将首先被 用标准顺序提取法从UMR-106细胞中提取 协议。然后使用DNA亲和层析分离NMP4和 COLA1的TH-1574至-1555个核苷酸片段的双链低聚物 启动子(B位)，其连接到交联化的琼脂糖基载体上。之后 洗脱后，NMP4将通过SDS/PAGE进一步纯化，提取并 用于抗体生产的微测序。对此的探测的可用性 核基质蛋白有助于进一步研究核基质蛋白的作用 NMP4在介导甲状旁腺激素作用的新的核途径中的作用，2)描绘了 甲状旁腺激素对骨的体内外作用差异 3)阐明甲状旁腺素合成代谢作用的细胞基础 在骨头上。

DESCRIPTION: The tissue matrix is the dynamic front that links the skeletal networks of the nucleus and the cytoplasm with the extracellular matrix. These studies are designed to test the hypothesis that nuclear matrix protein 4 (NMP4) a novel, PTH-responsive osteoblast nuclear matrix factor, plays a role in promoting expression of the alpha 1 chain of type 1 collagen (COLA1) in osteoblasts. The principal aim of these studies is to purify NMP4 for the purpose of obtaining microsequencing data needed to generate antibodies to thi novel protein. Nuclear matrix proteins will first be extracted from UMR-106 cells using a standard sequential extraction protocol. NMP4 will then be isolated using DNA-affinity chromatography and double-stranded oligomers to th -1574 to -1555 nt segment of the COLA1 promoter (site B), that is coupled to a crosslinked agarose support. After elution, NMP4 will be further purified by SDS/PAGE, excised and microsequenced for antibody production. The availability of probes to this nuclear matrix protein will 1) facilitate further studies on the role of NMP4 in mediating a novel nuclear pathway for PTH action, 2) delineate the differences between the in vitro and in vivo effects of PTH on bone formation, and 3) clarify the cellular basis for the anabolic actions of PTH on bone.