Deibel M R, Coleman M S
J Biol Chem. 1979 Sep 10;254(17):8634-40.
Terminal deoxynucleotidyltransferase has been purified from lymphoblasts of leukemic patients. The enzyme has a molecular weight of approximately 62,000 as determined by gel filtration and nondenaturing gel electrophoresis and is not dissociated into subunits by sodium dodecyl sulfate. In contrast, the terminal transferase enzyme from calf thymus has a molecular weight of 42,000 as determined by gel filtration, and is dissociated into 2 subunits of Mr 30,000 and 8,000 by sodium dodecyl sulfate. The enzyme has an isoelectric point of 8.2 and kinetic characteristics which are similar to those of calf thymus terminal transferase. The apparent Km for purine nucleotide polymerization at saturating initiator concentration with Mg2+ is 0.2 mM and with Mn2+ is 0.05 mM. Like calf terminal transferase, the reaction velocity is higher in the presence of Mg2+ than Mn2+. ATP inhibits the reaction catalyzed by terminal transferase isolated from human lymphoblasts due to mutual recognition of ATP and dATP by a common site on the enzyme. Preliminary experiments indicate that human terminal transferase may contain a small amount of carbohydrate. This report represents the first purification to near homogeneity of terminal transferase from a tissue source other than calf thymus.
末端脱氧核苷酸转移酶已从白血病患者的淋巴母细胞中纯化出来。通过凝胶过滤和非变性凝胶电泳测定，该酶的分子量约为62,000，并且不会被十二烷基硫酸钠解离成亚基。相比之下，通过凝胶过滤测定，来自小牛胸腺的末端转移酶分子量为42,000，并且会被十二烷基硫酸钠解离成分子量分别为30,000和8,000的两个亚基。该酶的等电点为8.2，其动力学特征与小牛胸腺末端转移酶相似。在饱和引发剂浓度下，Mg2+存在时嘌呤核苷酸聚合的表观Km为0.2 mM，Mn2+存在时为0.05 mM。与小牛末端转移酶一样，Mg2+存在时的反应速度高于Mn2+。由于酶上的一个共同位点对ATP和dATP的相互识别，ATP会抑制从人淋巴母细胞中分离出的末端转移酶催化的反应。初步实验表明，人末端转移酶可能含有少量碳水化合物。本报告首次报道了从小牛胸腺以外的组织来源将末端转移酶纯化至接近均一状态。
