Department of Biochemistry and Molecular Biology, University of Florida, Gainesville, FL, USA.
Mutagenesis. 2020 Dec 1;35(5):391-404. doi: 10.1093/mutage/geaa023.
DNA ligase I (LIG1) joins DNA strand breaks during DNA replication and repair transactions and contributes to genome integrity. The mutations (P529L, E566K, R641L and R771W) in LIG1 gene are described in patients with LIG1-deficiency syndrome that exhibit immunodeficiency. LIG1 senses 3'-DNA ends with a mismatch or oxidative DNA base inserted by a repair DNA polymerase. However, the ligation efficiency of the LIG1 variants for DNA polymerase-promoted mutagenesis products with 3'-DNA mismatches or 8-oxo-2'-deoxyguanosine (8-oxodG) remains undefined. Here, we report that R641L and R771W fail in the ligation of nicked DNA with 3'-8-oxodG, leading to an accumulation of 5'-AMP-DNA intermediates in vitro. Moreover, we found that the presence of all possible 12 non-canonical base pairs variously impacts the ligation efficiency by P529L and R771W depending on the architecture at the DNA end, whereas E566K exhibits no activity against all substrates tested. Our results contribute to the understanding of the substrate specificity and mismatch discrimination of LIG1 for mutagenic repair intermediates and the effect of non-synonymous mutations on ligase fidelity.
DNA 连接酶 I（LIG1）在 DNA 复制和修复过程中连接 DNA 链断裂，有助于基因组完整性。LIG1 基因中的突变（P529L、E566K、R641L 和 R771W）描述于存在免疫缺陷的 LIG1 缺陷综合征患者中。LIG1 通过修复 DNA 聚合酶检测具有错配或氧化 DNA 碱基插入的 3'-DNA 末端。然而，LIG1 变体对具有 3'-DNA 错配或 8-氧代-2'-脱氧鸟苷（8-oxodG）的 DNA 聚合酶促进的诱变产物的连接效率仍未定义。在这里，我们报告 R641L 和 R771W 无法连接带有 3'-8-oxodG 的缺口 DNA，导致体外积累 5'-AMP-DNA 中间体。此外，我们发现所有可能的 12 种非 canonical 碱基对根据 DNA 末端的结构，以不同的方式影响 P529L 和 R771W 的连接效率，而 E566K 对所有测试的底物均无活性。我们的研究结果有助于理解 LIG1 对诱变修复中间体的底物特异性和错配识别，以及非同义突变对连接酶保真度的影响。
