Lbvova T N, Tatarskaia R I
Biokhimiia. 1976 Aug;41(8):1426-34.
Unusual kinetics of the hydrolysis of the baker's yeast total tRNA and tRNA1Val by exonuclease A5 was found (two stages of the reaction; low initial velocity and still lower velocity of the second reaction stage; high Km values). These peculiarities were shown to be due to the three-dimensional structure of the substrate, which makes a large part of the tRNA molecule resistant to the exonuclease A5 attack. The exhaustive tRNA hydrolysis observed in the presence of very large doses of exonuclease A5 may be explained by a slight (less than 1%) endo-RNAase contamination in the exonuclease A5 preparation. The hydrolysis conditions become optimal, i.e. the exhaustive hydrolysis proceeds most rapidly, when the amount of endo-RNAase admixture reaches 20--30%. The artificial enzyme mixture of such a composition is a convenient reagent for RNA and expecially for tRNA hydrolysis under mild conditions to 5'-mononucleotides. The considerable resistance of tRNA to pure exonuclease A5 action will make it possible porbably to use tRNA instead of circular nucleic acids for endonuclease admixtures estimation in enzyme preparations.
发现核酸外切酶A5对面包酵母总tRNA和tRNA1Val的水解呈现出异常的动力学特征（反应分两个阶段；初始速度较低，第二反应阶段速度更低；米氏常数较高）。这些特性被证明是由于底物的三维结构所致，该结构使得tRNA分子的大部分对核酸外切酶A5的攻击具有抗性。在存在非常大量的核酸外切酶A5时观察到的tRNA彻底水解，可能是由于核酸外切酶A5制剂中存在少量（小于1%）的内切核糖核酸酶污染。当内切核糖核酸酶混合物的量达到20% - 30%时，水解条件变得最佳，即彻底水解进行得最快。这种组成的人工酶混合物是在温和条件下水解RNA尤其是tRNA为5'-单核苷酸的方便试剂。tRNA对纯核酸外切酶A5作用具有相当的抗性，这可能使得在酶制剂中用tRNA代替环状核酸来估计内切酶混合物成为可能。
