当前位置: X-MOL 学术Nat. Commun. › 论文详情
Our official English website, www.x-mol.net, welcomes your feedback! (Note: you will need to create a separate account there.)
Mechanism of substrate hydrolysis by the human nucleotide pool sanitiser DNPH1
Nature Communications ( IF 14.7 ) Pub Date : 2023-10-26 , DOI: 10.1038/s41467-023-42544-4
Neil J Rzechorzek 1 , Simone Kunzelmann 2 , Andrew G Purkiss 2 , Mariana Silva Dos Santos 3 , James I MacRae 3 , Ian A Taylor 4 , Kasper Fugger 1, 5 , Stephen C West 1
Affiliation  

Poly(ADP-ribose) polymerase (PARP) inhibitors are used in the clinic to treat BRCA-deficient breast, ovarian and prostate cancers. As their efficacy is potentiated by loss of the nucleotide salvage factor DNPH1 there is considerable interest in the development of highly specific small molecule DNPH1 inhibitors. Here, we present X-ray crystal structures of dimeric DNPH1 bound to its substrate hydroxymethyl deoxyuridine monophosphate (hmdUMP). Direct interaction with the hydroxymethyl group is important for substrate positioning, while conserved residues surrounding the base facilitate target discrimination. Glycosidic bond cleavage is driven by a conserved catalytic triad and proceeds via a two-step mechanism involving formation and subsequent disruption of a covalent glycosyl-enzyme intermediate. Mutation of a previously uncharacterised yet conserved glutamate traps the intermediate in the active site, demonstrating its role in the hydrolytic step. These observations define the enzyme’s catalytic site and mechanism of hydrolysis, and provide important insights for inhibitor discovery.



中文翻译:


人核苷酸池消毒剂 DNPH1 水解底物的机制



聚 (ADP-核糖) 聚合酶 (PARP) 抑制剂在临床上用于治疗 BRCA 缺陷型乳腺癌、卵巢癌和前列腺癌。由于核苷酸挽救因子 DNPH1 的缺失会增强它们的疗效,因此对开发高度特异性的小分子 DNPH1 抑制剂产生了相当大的兴趣。在这里,我们展示了与其底物羟甲基脱氧尿苷一磷酸 (hmdUMP) 结合的二聚体 DNPH1 的 X 射线晶体结构。与羟甲基基团的直接相互作用对于底物定位很重要,而碱基周围的保守残基有助于靶标区分。糖苷键裂解由保守的催化三联体驱动,并通过两步机制进行,涉及共价糖基-酶中间体的形成和随后的破坏。先前未表征但保守的谷氨酸的突变将中间体捕获在活性位点,证明其在水解步骤中的作用。这些观察结果确定了酶的催化位点和水解机制,并为抑制剂的发现提供了重要的见解。

更新日期:2023-10-26
down
wechat
bug