Keohavong P, Shukla R, Melacrinos A, Day B W, Reha-Krantz L
Department of Environmental and Occupational Health, University of Pittsburgh Cancer Institute, University of Pittsburgh, PA 15238, USA.
DNA Cell Biol. 1998 Jun;17(6):541-9. doi: 10.1089/dna.1998.17.541.
In vitro DNA replication by exonuclease-deficient T7 DNA polymerase (Sequenase) and an exonuclease deficient T4 DNA polymerase was examined on a 244-nucleotide DNA template treated with three electrophilic polycyclic aromatic hydrocarbon (PAH) metabolites: racemic trans-7,8-dihydroxy-anti-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BaPDE), trans-2,3-dihydroxy-anti-1,10b-epoxy-10b,1,2,3-tetrahydrofluoranthene (FADE), or 3,4-epoxy-3,4-dihydrocyclopenta[cd]pyrene (CPPE). The DNA replication terminated opposite template guanines and, to a lesser extent, at template adenines, as expected, as purines were modified preferentially by the chemical treatments. Analysis of the products synthesized on the damaged templates indicated that bypass replication by Sequenase proceeded in three steps: (1) replication first terminated one base 3' to each adduct; (2) a nucleotide was then incorporated opposite the PAH-modified base; and (3) replication continued at some sites to give full bypass of the lesions. The rate of lesion bypass was affected by the type of chemical adduct, the sequence context of the adduct, and the concentration of deoxynucleoside triphosphates. Short DNA repeats appeared to facilitate translesion replication.
利用缺乏核酸外切酶的T7 DNA聚合酶（测序酶）和缺乏核酸外切酶的T4 DNA聚合酶，在一个244个核苷酸的DNA模板上检测了体外DNA复制情况。该模板用三种亲电多环芳烃（PAH）代谢物进行了处理：外消旋反式-7,8-二羟基-反式-9,10-环氧-7,8,9,10-四氢苯并[a]芘（BaPDE）、反式-2,3-二羟基-反式-1,10b-环氧-10b,1,2,3-四氢荧蒽（FADE）或3,4-环氧-3,4-二氢环戊[cd]芘（CPPE）。正如预期的那样，DNA复制在模板鸟嘌呤相对处终止，在较小程度上也在模板腺嘌呤处终止，因为嘌呤在化学处理中优先被修饰。对在受损模板上合成的产物进行分析表明，测序酶的跨损伤复制分三步进行：（1）复制首先在每个加合物的3'端一个碱基处终止；（2）然后在PAH修饰的碱基相对处掺入一个核苷酸；（3）在一些位点复制继续进行，从而完全绕过损伤。损伤绕过的速率受化学加合物的类型、加合物的序列背景以及脱氧核苷三磷酸的浓度影响。短的DNA重复序列似乎有助于跨损伤复制。
