Kemp J D, Sutton D W, Hack E
Biochemistry. 1979 Aug 21;18(17):3755-60. doi: 10.1021/bi00584a017.
Nopaline synthase of sunflower (Helianthus annuus L.) crown gall tissue induced by Agrobacterium tumefaciens strain C58 or T37 (nopaline utilizers) was purified to homogeneity as judged by analytical disc gel electrophoresis. The native enzyme elutes from a column of Ultrogen AcA 34 as a single peak with an estimated molecular weight of 158,000. The dissociated enzyme migrates on NaDodSO4-polyacrylamide gels as a single band with a molecular weight of 40,000. Thus, the native enzyme appears to be composed of four equal-weight subunits. Nopaline synthesizing activity is found exclusively in crown gall tissues induced by strains of A. tumefaciens that utilize nopaline (e.g., C58 and T37). We found the same tissue specificity for the purified protein that we believe represents nopaline synthase. The results of kinetic studies of the purified enzyme are consistent with a ter-bi rapid-equilibrium random-order mechanism. Nopaline synthase is probably responsible for the in vivo synthesis of both N2-(1,3-dicarboxypropyl)arginine (nopaline) and N2-(1,3-dicarboxypropyl)ornithine (ornaline) in crown gall tissues since substrate specificities and Km values do not change during purification.
用根癌土壤杆菌C58或T37菌株（胭脂碱利用型）诱导向日葵（Helianthus annuus L.）冠瘿组织产生的胭脂碱合酶，经分析圆盘凝胶电泳鉴定已纯化至均一。天然酶从Ultrogen AcA 34柱上洗脱下来为单一峰，估计分子量为158,000。解离后的酶在NaDodSO4-聚丙烯酰胺凝胶上迁移为一条分子量为40,000的带。因此，天然酶似乎由四个等重亚基组成。胭脂碱合成活性仅在由利用胭脂碱的根癌土壤杆菌菌株（如C58和T37）诱导的冠瘿组织中发现。我们发现纯化后的蛋白质具有相同的组织特异性，我们认为该蛋白质代表胭脂碱合酶。对纯化酶的动力学研究结果与三底物快速平衡随机顺序机制一致。由于底物特异性和Km值在纯化过程中不变，胭脂碱合酶可能负责冠瘿组织中N2-(1,3-二羧丙基)精氨酸（胭脂碱）和N2-(1,3-二羧丙基)鸟氨酸（鸟氨酸）的体内合成。
