Yang Chien-Chin, Tseng Shu-Min, Pan Hung-Yin, Huang Chih-Hung, Chen Carton W
Department of Chemistry, Chung-Yuan Christian University, Chung-li, Taiwan.
Institute of Biochemical and Biomedical Engineering, National Taipei University of Technology, Taipei, Taiwan.
Nucleic Acids Res. 2017 Jun 2;45(10):5838-5849. doi: 10.1093/nar/gkx189.
Replication of the linear chromosomes of soil bacteria Streptomyces proceeds from an internal origin towards the telomeres, followed by patching of the resulting terminal single-strand overhangs by DNA synthesis using terminal proteins as the primer, which remains covalently bound to the 5΄ ends of the DNA. In most Streptomyces chromosomes, the end patching requires the single-strand overhangs, terminal protein Tpg, and terminal associated protein Tap. The telomere overhangs contain several palindromic sequences capable of forming stable hairpins. Previous in vitro deoxynucleotidylation studies indicated that Tap adds the Palindrome I sequence to Tpg, which is extended by a polymerase to fill the gap. In this study, the stringency of Palindrome I sequence was examined by an in vitro deoxynucleotidylation system and in vivo replication. Several nt in Palindrome I were identified to be critical for priming. While the first 3 G on the template were required for deoxynucleotidylation in vitro, deletions of them could be suppressed by the presence of dGTP. In vivo, deletions of these G were also tolerated, and the telomere sequence was restored in the linear plasmid DNA. Our results indicated that the truncated telomeres were repaired by extension synthesis by Tap on the foldback Palindrome I sequence.
土壤细菌链霉菌的线性染色体复制从内部起始点向端粒进行，随后通过以末端蛋白为引物进行DNA合成来填补产生的末端单链悬端，末端蛋白仍与DNA的5΄端共价结合。在大多数链霉菌染色体中，末端填补需要单链悬端、末端蛋白Tpg和末端相关蛋白Tap。端粒悬端包含几个能够形成稳定发夹结构的回文序列。先前的体外脱氧核苷酸化研究表明，Tap将回文I序列添加到Tpg上，然后由聚合酶将其延伸以填补缺口。在本研究中，通过体外脱氧核苷酸化系统和体内复制来检测回文I序列的严格性。确定回文I中的几个核苷酸对引发至关重要。虽然模板上的前3个G对于体外脱氧核苷酸化是必需的，但它们的缺失可被dGTP的存在所抑制。在体内，这些G的缺失也可被容忍，并且线性质粒DNA中的端粒序列得以恢复。我们的结果表明，截短的端粒通过Tap在回文I序列的回折上进行延伸合成来修复。
