Su T S, Beaudet A L, O'Brien W E
Biochemistry. 1981 May 12;20(10):2956-60. doi: 10.1021/bi00513a037.
The level of argininosuccinate synthetase activity in the human tissue culture cell line RPMI 2650 was 6-fold higher when citrulline was substituted for arginine in the culture medium. Canavanine-resistant (Canr) variants were isolated and had enzyme activity up to 25 nmol min-1 (mg of protein)-1 or 180-fold higher than that of the wild-type cells grown in arginine. The differences in enzyme activity were paralleled by differences in the amount of enzyme determined immunologically. The micrograms of enzyme per milligrams of protein, determined by complement fixation, were 0.03 for wild-type cells grown in arginine, 0.29 for wild-type cells grown in citrulline, and 6.73 for a Canr variant. In vivo labeling studies suggested increased synthesis of argininosuccinate synthetase in Canr cells, and in vitro translation of poly(adenylic acid) [poly(A)] messenger ribonucleic acid (mRNA) from wild-type and Canr cells confirmed a quantitatively compatible increase in translatable poly(A) mRNA for the enzyme in Canr cells. No precursor for the enzyme was recognized by using in vitro translation, and the poly(A) mRNA for the enzyme had a sedimentation value of 16 S by sucrose-gradient analysis. The levels of argininosuccinate synthetase activity in the Canr cells were similar to those found in normal liver.
当在培养基中用瓜氨酸替代精氨酸时，人组织培养细胞系RPMI 2650中的精氨琥珀酸合成酶活性水平提高了6倍。分离出了抗刀豆氨酸（Canr）变体，其酶活性高达25 nmol·min⁻¹·（mg蛋白质）⁻¹，比在精氨酸中生长的野生型细胞高180倍。酶活性的差异与通过免疫方法测定的酶量差异平行。通过补体固定法测定，每毫克蛋白质中酶的微克数，在精氨酸中生长的野生型细胞为0.03，在瓜氨酸中生长的野生型细胞为0.29，而一个Canr变体为6.73。体内标记研究表明Canr细胞中精氨琥珀酸合成酶的合成增加，对野生型和Canr细胞的聚腺苷酸[poly(A)]信使核糖核酸（mRNA）进行体外翻译，证实Canr细胞中该酶的可翻译poly(A) mRNA在数量上有相应增加。通过体外翻译未识别出该酶的前体，通过蔗糖梯度分析，该酶的poly(A) mRNA沉降值为16 S。Canr细胞中精氨琥珀酸合成酶的活性水平与正常肝脏中的相似。
