DIRECT IDENTIFICATION OF CALCIUM-BINDING AMINO-ACID, GAMMA-CARBOXYGLUTAMATE, IN MINERALIZED TISSUE
DIRECT IDENTIFICATION OF CALCIUM-BINDING AMINO-ACID, GAMMA-CARBOXYGLUTAMATE, IN MINERALIZED TISSUE
复制标题
DOI：
10.1073/pnas.72.10.3925
复制
发表时间：
1975-01-01
影响因子：
11.1
通讯作者：
GALLOP, PM
中科院分区：
文献类型：
--
作者：
HAUSCHKA, PV;LIAN, JB;GALLOP, PM
已经开发出一种直接方法来定量鉴定蛋白质中的钙结合氨基酸γ-羧基谷氨酸。这应该是有利的许多系统的研究，其中钙或其他二价阳离子的结合的具体作用被怀疑。矿化组织的调查，其中钙结合蛋白参与矿化过程中，发现γ-羧基谷氨酸是存在于蛋白质溶解从鸡骨与中性水溶液乙二胺四乙酸。这是通过从碱性水解产物中直接分离氨基酸并通过在100 ℃下在0.05 M HCl中脱羧将其定量转化为谷氨酸而建立的。脱羧动力学和色谱行为与人凝血酶原中γ-羧基谷氨酸的脱羧动力学和色谱行为相同。通过磷酸盐梯度洗脱从羟基磷灰石中分离可溶性骨蛋白后，发现γ-羧基谷氨酸主要集中在一种BaSO 4可吸附阴离子蛋白质中;骨胶原缺乏氨基酸。鉴于最近发现的维生素K的需要产生的钙结合位点（γ-羧基谷氨酸）的特定谷氨酸残基在凝血酶原的γ-羧化，我们的研究结果可能涉及维生素K的代谢在正常的骨发育和建议的作用，γ-羧基谷氨酸丰富的蛋白质在调节钙盐沉积在矿化组织。
A direct approach has been developed for quantitative identification of the calcium-binding amino acid, gamma-carboxyglutamate, in proteins. This should be advantageous for the study of numerous systems where specific roles for the binding of calcium or other divalent cations are suspected. Investigation of mineralized tissue, where calcium-binding proteins are implicated in the mineralization process, revealed that gamma-carboxyglutamate was present in proteins solubilized from chicken bone with neutral aqueous ethylenediamine tetraacetic acid. This was established by direct isolation of the amino acid from alkaline hydrolysates and its quantitative conversion to glutamic acid by decarboxylation in 0.05 M HCl at 100 degrees. The kinetics of decarboxylation and chromatographic behavior are identical to those of gamma-carboxyglutamate from human prothrombin. After resolution of the soluble bone proteins by phosphate gradient elution from hydroxyapatite, gamma-carboxyglutamate was found to be concentrated primarily in one BaSO4-adsorbable anionic protein species; bone collagen was devoid of the amino acid. In view of the recently discovered requirement of vitamin K for generation of calcium binding sites (gamma-carboxyglutamate) by gamma-carboxylation of specific glutamic acid residues in prothrombin, our findings may implicate vitamin K metabolism in normal bone development and suggest a role for the gamma-carboxyglutamate-rich protein in regulation of calcium salt deposition in mineralized tissues.