High-affinity interaction of poly(ADP-ribose) and the human DEK oncoprotein depends upon chain length.
High-affinity interaction of poly(ADP-ribose) and the human DEK oncoprotein depends upon chain length.
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DOI：
10.1021/bi1004365
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发表时间：
2010-08-24
期刊：
影响因子：
2.9
通讯作者：
Kappes, Ferdinand
中科院分区：
文献类型：
--
作者：
Fahrer, Joerg;Popp, Oliver;Malanga, Maria;Beneke, Sascha;Markovitz, David M.;Ferrando-May, Elisa;Buerkle, Alexander;Kappes, Ferdinand
聚腺苷二磷酸核糖聚合酶-1（Poly（ADP-ribose）polymerase-1，PARP-1）是一种分子DNA损伤传感器，在NAD+的作用下催化合成复杂的生物聚合物聚腺苷二磷酸核糖[poly（ADP-ribose）[PAR]。PAR参与基本的细胞过程，如DNA代谢和转录，并与参与DNA修复和染色质结构调节的特异性结合蛋白非共价相互作用。与DNA修复和染色质组织有关的一个因子是DEK癌蛋白，它是后生动物染色质的一种丰富和保守的成分，也是其蛋白质类的唯一成员。我们最近已经证明，DEK，在应力条件下，共价修饰PAR的PARP-1，导致部分释放到细胞质中的DEK。此外，我们还观察到DEK和PAR之间的非共价相互作用，我们在本工作中详细介绍。使用序列比对，我们在DEK一级序列中鉴定了三个功能性PAR结合位点，并确认了它们在PAR结合研究中的功能。此外，我们表明，非共价结合DEK是依赖于PAR链的长度所揭示的重叠印迹技术和PAR EMSA。有趣的是，DEK促进与54聚体PAR（KD=6 × 10−8 M）形成确定的复合物，而与短PAR链（18聚体）没有检测到特异性相互作用。与DEK的共价聚（ADP-核糖基）化形成鲜明对比，非共价相互作用不影响DEK结合DNA的总体能力。相反，非共价相互作用干扰DEK随后的DNA依赖性多聚化活性，如在South-Western、EMSA、拓扑学和聚集测定中所见。特别地，PAR与DEK的非共价连接通过与DNA结合竞争促进DEK-DEK复合物的形成。这可以通过PAR结合的DEK对溶液中DNA模板的亲和力降低来观察。两者合计，我们的研究结果加深了分子的DEK-PAR相互作用的理解，并支持存在的细胞“PAR代码”的PAR链长度表示。
Poly(ADP-ribose) polymerase-1 (PARP-1) is a molecular DNA damage sensor that catalyzes the synthesis of the complex biopolymer poly(ADP-ribose) [PAR] under consumption of NAD+. PAR engages in fundamental cellular processes such as DNA metabolism and transcription, and interacts non-covalently with specific binding proteins involved in DNA repair and regulation of chromatin structure. A factor implicated in DNA repair and chromatin organization is the DEK oncoprotein, an abundant and conserved constituent of metazoan chromatin, and the only member of its protein class. We have recently demonstrated that DEK, under stress conditions, is covalently modified with PAR by PARP-1, leading to a partial release of DEK into the cytoplasm. Additionally, we have also observed a non-covalent interaction between DEK and PAR, which we detail in the present work. Using sequence alignment, we identify three functional PAR-binding sites in the DEK primary sequence and confirm their functionality in PAR binding studies. Furthermore, we show that the non-covalent binding to DEK is dependent on PAR chain length as revealed by an overlay blot technique and PAR EMSA. Intriguingly, DEK promotes the formation of a defined complex with a 54mer PAR (KD=6 × 10−8 M), whereas no specific interaction is detected with a short PAR chain (18mer). In stark contrast to covalent poly(ADP-ribosyl)ation of DEK, the non-covalent interaction does not affect the overall ability of DEK to bind to DNA. Instead the non-covalent interaction interferes with subsequent DNA-dependent multimerization activities of DEK, as seen in South-Western, EMSA, topology and aggregation assays. In particular, non-covalent attachment of PAR to DEK promotes the formation of DEK-DEK complexes by competing with DNA binding. This was seen by the reduced affinity of PAR-bound DEK for DNA templates in solution. Taken together, our findings deepen the molecular understanding of the DEK-PAR interplay and support the existence of a cellular “PAR code” represented by PAR chain length.
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影响因子：
3.8
作者：
Goodarzi, AA;Lees-Miller, SP
通讯作者：
Lees-Miller, SP
影响因子：
2.9
作者：
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通讯作者：
Zhang, J
影响因子：
4
作者：
Ame, Jean-Christophe;Fouquerel, Elise;Schreiber, Valerie
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影响因子：
64.8
作者：
Ahel, Ivan;Ahel, Dragana;West, Stephen C.
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West, Stephen C.
DOI：
10.1016/s0006-291x(87)80219-x
发表时间：
1987-11-13
影响因子：
3.1
作者：
GRADWOHL, G;MAZEN, A;DEMURCIA, G
通讯作者：
DEMURCIA, G