The Rev1 interacting region (RIR) motif in the scaffold protein XRCC1 mediates a low-affinity interaction with polynucleotide kinase/phosphatase (PNKP) during DNA single-strand break repair.
The Rev1 interacting region (RIR) motif in the scaffold protein XRCC1 mediates a low-affinity interaction with polynucleotide kinase/phosphatase (PNKP) during DNA single-strand break repair.
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DOI：
10.1074/jbc.m117.806638
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发表时间：
2017-09-29
期刊：
影响因子：
--
通讯作者：
Caldecott KW
中科院分区：
文献类型：
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作者：
Breslin C;Mani RS;Fanta M;Hoch N;Weinfeld M;Caldecott KW
支架蛋白X射线修复交叉互补1（XRCC 1）与参与DNA碱基切除修复和单链断裂修复（SSBR）的多种酶相互作用，对遗传完整性和正常神经功能非常重要。XRCC 1最重要的相互作用之一是与多核苷酸激酶/磷酸酶（PNKP）的相互作用，PNKP是一种双功能DNA激酶/磷酸酶，可处理受损的DNA末端，如果发生突变，则会导致共济失调伴眼神经失用症4（AOA 4）和小头畸形伴早发性癫痫发作和发育迟缓（MCSZ）。XRCC 1和PNKP通过XRCC 1中的高亲和力磷酸化依赖性相互作用位点和PNKP中的叉头相关结构域相互作用。在这里，我们使用生物化学和生物物理方法鉴定了XRCC 1中的第二个PNKP相互作用位点，该位点以较低的亲和力结合PNKP，并且独立于XRCC 1磷酸化。然而，这种相互作用仍然刺激PNKP活性，并促进SSBR和细胞存活。低亲和力相互作用位点需要XRCC 1中高度保守的Rev 1相互作用区（RIR）基序，并包括三个关键的和进化上不变的苯丙氨酸残基。我们提出了一个二分相互作用模型，其中先前确定的高亲和力相互作用作为分子系链，将XRCC 1和PNKP保持在一起，从而促进这里确定的低亲和力相互作用，然后直接刺激PNKP。
The scaffold protein X-ray repair cross-complementing 1 (XRCC1) interacts with multiple enzymes involved in DNA base excision repair and single-strand break repair (SSBR) and is important for genetic integrity and normal neurological function. One of the most important interactions of XRCC1 is that with polynucleotide kinase/phosphatase (PNKP), a dual-function DNA kinase/phosphatase that processes damaged DNA termini and that, if mutated, results in ataxia with oculomotor apraxia 4 (AOA4) and microcephaly with early-onset seizures and developmental delay (MCSZ). XRCC1 and PNKP interact via a high-affinity phosphorylation-dependent interaction site in XRCC1 and a forkhead-associated domain in PNKP. Here, we identified using biochemical and biophysical approaches a second PNKP interaction site in XRCC1 that binds PNKP with lower affinity and independently of XRCC1 phosphorylation. However, this interaction nevertheless stimulated PNKP activity and promoted SSBR and cell survival. The low-affinity interaction site required the highly conserved Rev1-interacting region (RIR) motif in XRCC1 and included three critical and evolutionarily invariant phenylalanine residues. We propose a bipartite interaction model in which the previously identified high-affinity interaction acts as a molecular tether, holding XRCC1 and PNKP together and thereby promoting the low-affinity interaction identified here, which then stimulates PNKP directly.