Ogawa H, Goldsmith L A
J Biol Chem. 1976 Dec 10;251(23):7281-8.
A transglutaminase from human hair follicle-free epidermis was purified to homogeneity using gel filtration and ion exchange chromatography. The enzyme had an apparent Mr = 51,000 +/- 2,000 by sodium dodecyl sulfate electrophoresis, 100,000 +/- 5,000 by discontinuous gel electrophoresis, and 50,000 +/- 2,000 by gel filtration in Bio-Gel A-0.5m agarose. The enzyme cross-linked Factor XIII-free fibrinogen forming gamma dimers and alpha polymers. Either calcium or strontium was necessary for enzyme activity. In the presence of calcium, enzyme activity was increased by heating at 56 degrees or by treating with dimethylsulfoxide. Activation required calcium and occurred in the presence of serine protease inhibitors. The activated and native enzyme had apparently identical mobilities in acrylamide disc electrophoresis and sodium dodecyl sulfate electrophoresis. The Km values for two substrates in the reaction, casein and putrescine, were very similar for the native and the activated enzyme. The activated enzyme had a larger elution volume on Bio-Gel A-0.5m in the presence of calcium than did the native enzyme. The detailed mechanism of activation remains to be determined.
利用凝胶过滤和离子交换色谱法，从不含人毛囊的表皮中纯化出一种转谷氨酰胺酶，使其达到均一状态。通过十二烷基硫酸钠电泳，该酶的表观分子量为51,000 ± 2,000；通过不连续凝胶电泳，其表观分子量为100,000 ± 5,000；在Bio-Gel A-0.5m琼脂糖中进行凝胶过滤时，其表观分子量为50,000 ± 2,000。该酶可使不含因子XIII的纤维蛋白原交联形成γ二聚体和α聚合物。钙或锶对于酶活性是必需的。在有钙存在的情况下，通过56℃加热或用二甲基亚砜处理可提高酶活性。激活需要钙，且在丝氨酸蛋白酶抑制剂存在时发生。在丙烯酰胺圆盘电泳和十二烷基硫酸钠电泳中，活化酶和天然酶的迁移率明显相同。对于该反应中的两种底物酪蛋白和腐胺，天然酶和活化酶的米氏常数非常相似。在有钙存在的情况下，活化酶在Bio-Gel A-0.5m上的洗脱体积比天然酶大。激活的详细机制仍有待确定。
