Kato M, Nozawa Y
Anal Biochem. 1984 Dec;143(2):361-7. doi: 10.1016/0003-2697(84)90675-4.
Arginyltransferase was purified 60,000-fold from the 105,000g supernatant of hog kidney by fractionation with ammonium sulfate, precipitation at pH 5.2, and chromatographies on DEAE-cellulose and hydroxyapatite, as well as affinity chromatographies on heparin-Sepharose and angiotensin II-Sepharose. Purified transferase had a specific activity of 7.6 mumol/min/mg, which was 66 times higher than that attained thus far (R. L. Soffer, 1970, J. Biol. Chem. 245, 731-737). Electrophoresis of purified transferase on polyacrylamide gel showed a single band which corresponded to the region of activity in homogenized slices of a duplicate gel. The homogeneity was also observed by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, where only a single band of Mr 35,000 was observed, even after staining by the silver method (B. R. Oakley, D. R. Kirsch, and N. R. Morris, 1980, Anal. Biochem. 105, 361-363). Antibody to the purified transferase was prepared in a rabbit. The antibody inhibited arginyltransferase activity, and showed a single precipitin line by double immunodiffusion with crude transferase preparation.
通过硫酸铵分级分离、pH 5.2沉淀、DEAE - 纤维素和羟基磷灰石层析以及肝素 - 琼脂糖和血管紧张素II - 琼脂糖亲和层析，从猪肾105,000g上清液中纯化精氨酰转移酶60,000倍。纯化后的转移酶比活性为7.6 μmol/min/mg，比迄今为止所达到的比活性高66倍（R. L. Soffer，1970，《生物化学杂志》245，731 - 737）。纯化后的转移酶在聚丙烯酰胺凝胶上电泳显示出一条单一的条带，与重复凝胶匀浆切片中的活性区域相对应。在十二烷基硫酸钠存在下进行聚丙烯酰胺凝胶电泳时也观察到了均一性，即使采用银染法（B. R. Oakley、D. R. Kirsch和N. R. Morris，1980，《分析生物化学》105，361 - 363），也仅观察到一条Mr为35,000的单一谱带。用纯化后的转移酶在兔体内制备抗体。该抗体抑制精氨酰转移酶活性，并且与粗转移酶制剂进行双向免疫扩散时显示出一条单一的沉淀线。
