当前位置: X-MOL 学术RSC Chem. Biol. › 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.)
Site-specific modification and segmental isotope labelling of HMGN1 reveals long-range conformational perturbations caused by posttranslational modifications
RSC Chemical Biology Pub Date : 2021-1-5 , DOI: 10.1039/d0cb00175a
Gerhard Niederacher 1 , Debra Urwin 2 , Yasmin Dijkwel 2 , David J Tremethick 2 , K Johan Rosengren 3 , Christian F W Becker 1 , Anne C Conibear 3
Affiliation  

Interactions between histones, which package DNA in eukaryotes, and nuclear proteins such as the high mobility group nucleosome-binding protein HMGN1 are important for regulating access to DNA. HMGN1 is a highly charged and intrinsically disordered protein (IDP) that is modified at several sites by posttranslational modifications (PTMs) – acetylation, phosphorylation and ADP-ribosylation. These PTMs are thought to affect cellular localisation of HMGN1 and its ability to bind nucleosomes; however, little is known about how these PTMs regulate the structure and function of HMGN1 at a molecular level. Here, we combine the chemical biology tools of protein semi-synthesis and site-specific modification to generate a series of unique HMGN1 variants bearing precise PTMs at their N- or C-termini with segmental isotope labelling for NMR spectroscopy. With access to these precisely-defined variants, we show that PTMs in both the N- and C-termini cause changes in the chemical shifts and conformational populations in regions distant from the PTM sites; up to 50–60 residues upstream of the PTM site. The PTMs investigated had only minor effects on binding of HMGN1 to nucleosome core particles, suggesting that they have other regulatory roles. This study demonstrates the power of combining protein semi-synthesis for introduction of site-specific PTMs with segmental isotope labelling for structural biology, allowing us to understand the role of PTMs with atomic precision, from both structural and functional perspectives.

中文翻译:

HMGN1 的位点特异性修饰和片段同位素标记揭示了翻译后修饰引起的长程构象扰动

在真核生物中包装 DNA 的组蛋白与核蛋白(如高迁移率基团核小体结合蛋白 HMGN1)之间的相互作用对于调节 DNA 的获取非常重要。HMGN1 是一种高度带电且本质上无序的蛋白质 (IDP),它在多个位点通过翻译后修饰 (PTM) 进行修饰——乙酰化、磷酸化和 ADP 核糖基化。这些 PTM 被认为会影响 HMGN1 的细胞定位及其结合核小体的能力;然而,关于这些 PTM 如何在分子水平上调节 HMGN1 的结构和功能,我们知之甚少。在这里,我们结合了蛋白质半合成和位点特异性修饰的化学生物学工具,以生成一系列独特的 HMGN1 变体,这些变体在其 N 或 C 末端带有精确的 PTM,并带有用于 NMR 光谱的分段同位素标记。通过访问这些精确定义的变体,我们表明 N 端和 C 端的 PTM 会导致远离 PTM 位点的区域的化学位移和构象种群发生变化;PTM 位点上游多达 50-60 个残基。研究的 PTM 对 HMGN1 与核小体核心颗粒的结合只有很小的影响,表明它们具有其他调节作用。这项研究证明了结合蛋白质半合成引入位点特异性 PTM 与用于结构生物学的节段同位素标记的能力,使我们能够从结构和功能的角度以原子精度理解 PTM 的作用。我们表明 N 端和 C 端的 PTM 会导致远离 PTM 位点的区域的化学位移和构象群发生变化;PTM 位点上游多达 50-60 个残基。研究的 PTM 对 HMGN1 与核小体核心颗粒的结合只有很小的影响,表明它们具有其他调节作用。这项研究证明了结合蛋白质半合成引入位点特异性 PTM 与用于结构生物学的节段同位素标记的能力,使我们能够从结构和功能的角度以原子精度理解 PTM 的作用。我们表明 N 端和 C 端的 PTM 会导致远离 PTM 位点的区域的化学位移和构象群发生变化;PTM 位点上游多达 50-60 个残基。研究的 PTM 对 HMGN1 与核小体核心颗粒的结合只有很小的影响,表明它们具有其他调节作用。这项研究证明了结合蛋白质半合成引入位点特异性 PTM 与用于结构生物学的节段同位素标记的能力,使我们能够从结构和功能的角度以原子精度理解 PTM 的作用。
更新日期:2021-01-05
down
wechat
bug