Hughes J, Mellows G
Biochem J. 1980 Oct 1;191(1):209-19. doi: 10.1042/bj1910209.
Sodium pseudomonate was shown to be a powerful competitive inhibitor of Escherichia coli B isoleucyl-tRNA synthetase (Ile-tRNA synthetase). The antibiotic competitively inhibits (Ki 6 nM; cf. Km 6.3 microM), with respect top isoleucine, the formation of the enzyme . Ile approximately AMP complex as measured by the pyrophosphate-exchange reaction, and has no effect on the transfer of [14C]isoleucine from the enzyme . [14C]Ile approximately AMP complex to tRNAIle. The inhibitory constant for the pyrophosphate-exchange reaction was of the same order as that determined for the inhibition of the overall aminoacylation reaction (Ki 2.5 nM; cf. Km 11.1 microM). Sodium [9'-3H]pseudomonate forms a stable complex with Ile-tRNA synthetase. Gel-filtration and gel-electrophoresis studies showed that the antibiotic is only fully released from the complex by 5 M-urea treatment or boiling in 0.1% sodium dodecyl sulphate. The molar binding ratio of sodium [9'-3H]pseudomonate to Ile-tRNA synthetase was found to be 0.85:1 by equilibrium dialysis. Aminoacylation of yeast tRNAIle by rat liver Ile-tRNA synthetase was also competitively inhibited with respect to isoleucine, Ki 20 microM (cf. Km 5.4 microM). The Km values for the rat liver and E. coli B enzymes were of the same order, but the Ki for the rat liver enzyme was 8000 times the Ki for the E. coli B enzyme. This presumably explains the low toxicity of the antibiotic in mammals.
假单胞菌酸钠被证明是大肠杆菌B异亮氨酰 - tRNA合成酶（Ile - tRNA合成酶）的一种强效竞争性抑制剂。就异亮氨酸而言，该抗生素竞争性抑制（Ki为6 nM；对比Km为6.3 μM）酶 - Ile·AMP复合物的形成，这是通过焦磷酸交换反应测定的，并且对[14C]异亮氨酸从酶 - [14C]Ile·AMP复合物转移到tRNAIle没有影响。焦磷酸交换反应的抑制常数与对整体氨酰化反应抑制所测定的常数处于同一数量级（Ki为2.5 nM；对比Km为11.1 μM）。[9'-3H]假单胞菌酸钠与Ile - tRNA合成酶形成稳定复合物。凝胶过滤和凝胶电泳研究表明，只有通过5 M尿素处理或在0.1%十二烷基硫酸钠中煮沸，抗生素才会从复合物中完全释放出来。通过平衡透析发现，[9'-3H]假单胞菌酸钠与Ile - tRNA合成酶的摩尔结合比为0.85:1。大鼠肝脏Ile - tRNA合成酶对酵母tRNAIle的氨酰化作用在异亮氨酸方面也受到竞争性抑制，Ki为20 μM（对比Km为5.4 μM）。大鼠肝脏和大肠杆菌B酶的Km值处于同一数量级，但大鼠肝脏酶的Ki是大肠杆菌B酶的Ki的8000倍。这大概解释了该抗生素在哺乳动物中的低毒性。
