放线菌素D和三磷酸双脱氧胸苷对人细胞DNA合成多酶复合物的抑制作用。

Inhibition by aphidicolin and dideoxythymidine triphosphate of a multienzyme complex of DNA synthesis from human cells.

作者信息

Wickremasinghe R G, Hoffbrand A V

出版信息

FEBS Lett. 1983 Aug 8;159(1-2):175-9. doi: 10.1016/0014-5793(83)80441-4.

Abstract

A multienzyme complex consisting of DNA polymerase and several DNA precursor-synthesizing enzymes was solubilized by gentle lysis of cultured human cells. This complex channelled the distal precursor [3H]dTMP into DNA. The patterns of inhibition of the complex by aphidicolin and dideoxythymidine triphosphate (ddTTP) suggested that the complex contained the replicative DNA polymerase, polymerase alpha. Inhibition by ddTTP was competitive with dTTP. This was exploited to estimate the effective concentration of [3H]dTTP at the site of DNA synthesis during channelling of [3H]dTMP into DNA. The estimated concentration (about 50 microM) was so high as to suggest that the solubilized complex was able to functionally compartmentalize DNA precursors.

摘要

通过对培养的人类细胞进行温和裂解，可溶解一种由DNA聚合酶和几种DNA前体合成酶组成的多酶复合物。该复合物将远端前体[3H]dTMP导入DNA。放线菌素D和双脱氧胸苷三磷酸（ddTTP）对该复合物的抑制模式表明，该复合物含有复制性DNA聚合酶α。ddTTP的抑制作用与dTTP具有竞争性。利用这一点来估计在将[3H]dTMP导入DNA的过程中，DNA合成部位[3H]dTTP的有效浓度。估计浓度（约50微摩尔）如此之高，表明溶解的复合物能够在功能上分隔DNA前体。