Bell D E, Fridland A
Biochim Biophys Acta. 1980;606(1):57-66. doi: 10.1016/0005-2787(80)90097-0.
The effects of 9-beta-D-arabinosyladenine (AraAde), 1-beta-D-arabinosylcytosine (AraCyt) and 2'-deoxyadenosine on DNA replication in cultured human lymphoblasts (CCRF-CEM line) were studied by pulse-labeling cells with [3H]thymidine and analyzing the nascent DNA by velocity sedimentation in alkaline sucrose gradients. At doses that reduced the overall rate of DNA synthesis to 50--70% of control values, both AraAde and AraCyt profoundly inhibited the formation of new replicons, with secondary effects on chain elongation contributing to the total inhibition of DNA synthesis. In contrast, the suppression of DNA synthesis by 2'-deoxyadenosine stemmed mainly from an inhibition of chain elongation. These studies also disclosed that about 100 times more AraAde than AraCyt was required to produce a similar inhibition of DNA replication in CCRF-CEM cells. Determination of intracellular concentrations of the nucleoside triphosphates (AraCTP and AraATP) indicated to 90% inhibition of DNA synthesis was achieved at 1.6 and 25 pmol/1 . 10(6) cells, respectively. Studies with cell lysates revealed that the replicative machinery in CCRF-CEM cells is more sensitive to AraCTP than to AraATP. This finding contrasts with earlier research, in which the inhibtion of purified DNA polymerase by either AraATP of AraCTP yielded essentially the same Ki value. The difference in sensitivity of the cell lysate to these arabinonucleotides may reflect either a target enzyme other than DNA polymerase or, more plausibly, some subtle interaction of the polymerase with other components of the replicative process.
通过用[³H]胸腺嘧啶脉冲标记细胞，并在碱性蔗糖梯度中通过速度沉降分析新生DNA，研究了9-β-D-阿拉伯糖基腺嘌呤（AraAde）、1-β-D-阿拉伯糖基胞嘧啶（AraCyt）和2'-脱氧腺苷对培养的人淋巴母细胞（CCRF-CEM系）中DNA复制的影响。在将DNA合成的总体速率降低至对照值的50%-70%的剂量下，AraAde和AraCyt均显著抑制新复制子的形成，对链延长的次要影响导致了DNA合成的总体抑制。相比之下，2'-脱氧腺苷对DNA合成的抑制主要源于对链延长的抑制。这些研究还表明，在CCRF-CEM细胞中产生类似的DNA复制抑制所需的AraAde比AraCyt多约100倍。核苷三磷酸（AraCTP和AraATP）的细胞内浓度测定表明，分别在1.6和25 pmol/1.10⁶个细胞时实现了90%的DNA合成抑制。对细胞裂解物的研究表明，CCRF-CEM细胞中的复制机制对AraCTP比对AraATP更敏感。这一发现与早期研究形成对比，在早期研究中，AraATP或AraCTP对纯化的DNA聚合酶的抑制产生的Ki值基本相同。细胞裂解物对这些阿拉伯核苷酸敏感性的差异可能反映了除DNA聚合酶之外的靶酶，或者更合理地说，反映了聚合酶与复制过程中其他成分的一些微妙相互作用。
