Basnak'ian A G, Bubnov N V, Votrin I I
Biokhimiia. 1989 Feb;54(2):273-83.
In order to determine the ratio of activities of major endonucleases of rat liver chromatin, a stepwise fractionation of cell nuclear extracts by chromatography on phosphocellulose and gel filtration through Toyopearl HW60 was carried out. This procedure resulted in partially purified preparations of Ca2+,Mg2+-dependent endonuclease (55 +/- 10 kD), Ca2+,Mg2+-dependent endonuclease (30 +/- 10 kD), Mn2+-dependent endonuclease (30 +/- 5 kD) and acid cation-independent endonuclease. The Ca2+,Mg2+-dependent endonuclease with Mr of 55 +/- 10 kD made up to 57% of the nuclear extract activity in the presence of Ca2+ + Mg2+ and revealed a high calcium-magnesium synergism. Under the same experimental conditions, the 30 +/- 10 kD enzyme made up to 33% of the nuclear extract activity and revealed a low synergism. The activity of Mn2+-dependent endonuclease made up to 26% of the total nuclear extract activity in the presence of Mn2+, that of acid endonuclease--11% of the extract activity in 1 mM EDTA at pH 5.0. It was assumed that the low molecular weight Ca2+,Mg2+-dependent endonuclease represents a product of limited proteolysis of high molecular weight Ca2+,Mg2+-dependent endonuclease.
为了确定大鼠肝脏染色质主要核酸内切酶的活性比例，通过磷酸纤维素柱色谱和经Toyopearl HW60凝胶过滤对细胞核提取物进行分步分级分离。此方法得到了部分纯化的Ca2+、Mg2+依赖性核酸内切酶（55±10 kD）、Ca2+、Mg2+依赖性核酸内切酶（30±10 kD）、Mn2+依赖性核酸内切酶（30±5 kD）和酸性阳离子非依赖性核酸内切酶制剂。在Ca2+ + Mg2+存在下，分子量为55±10 kD的Ca2+、Mg2+依赖性核酸内切酶占核提取物活性的57%，并表现出高钙镁协同作用。在相同实验条件下，30±10 kD的酶占核提取物活性的33%，并表现出低协同作用。在Mn2+存在下，Mn2+依赖性核酸内切酶的活性占核提取物总活性的26%，酸性核酸内切酶的活性在pH 5.0的1 mM EDTA中占提取物活性的11%。据推测，低分子量的Ca2+、Mg2+依赖性核酸内切酶是高分子量Ca2+、Mg2+依赖性核酸内切酶有限蛋白水解的产物。
