The SARS-unique domain (SUD) of SARS coronavirus contains two macrodomains that bind G-quadruplexes.
The SARS-unique domain (SUD) of SARS coronavirus contains two macrodomains that bind G-quadruplexes.
复制标题
DOI：
10.1371/journal.ppat.1000428
复制
发表时间：
2009-05
期刊：
影响因子：
6.7
通讯作者：
Hilgenfeld R
中科院分区：
文献类型：
--
作者：
Tan J;Vonrhein C;Smart OS;Bricogne G;Bollati M;Kusov Y;Hansen G;Mesters JR;Schmidt CL;Hilgenfeld R
自2003年严重急性呼吸综合征（SARS）爆发以来，SARS冠状病毒（SARS-CoV）的复制酶/转录酶组分--非结构蛋白（Nsps）的三维结构已被确定。然而，在大Nsp3（1922个氨基酸残基），所谓的SARS独特结构域（SUD）的结构和功能仍然难以捉摸。SUD只发生在SARS-CoV和在某些蝙蝠中发现的高度相关的病毒中，但不存在于所有其他冠状病毒中。因此，有人推测，与其他冠状病毒相比，它可能参与了SARS-CoV的极端致病性，其中大多数冠状病毒仅引起人类轻度感染。为了帮助阐明SUD的功能，我们已经确定了Nsp 3的片段389 - 652（"SUDcore"）的晶体结构，其包含该结构域的338个残基中的264个。单斜和三斜晶型（分辨率分别为2.2和2.8 μ m）显示SUDcore形成同二聚体。每个单体由两个亚结构域SUD-N和SUD-M组成，其宏结构域折叠类似于SARS-CoV X结构域。然而，与后者相反，SUD不能结合ADP-核糖，如通过区域干扰凝胶电泳所确定的。相反，整个SUDcore以及它的各个亚结构域与已知形成G-四链体的寡核苷酸相互作用。这包括寡脱氧核苷酸以及寡核糖核苷酸。SUD-N亚结构域表面的赖氨酸残基的突变导致G-四链体结合的减少，而SUD-M亚结构域的突变则使其失效。由于没有证据表明Nsp3进入宿主细胞的核，因此SARS-CoV基因组RNA或含有长G-链段的宿主细胞mRNA可能是SUD的靶点。SARS-CoV基因组中没有超过5 - 6个核苷酸的G片段，但在编码某些参与细胞凋亡或信号转导的宿主细胞蛋白质的mRNA的3 ′非翻译区中发现了更多的G片段，并已被证明在体外与SUD结合。因此，SUD可能参与控制宿主细胞对病毒感染的反应。可能的干扰与聚（ADP-核糖）聚合酶样结构域进行了讨论。SARS冠状病毒的基因组编码16种非结构蛋白，这些蛋白参与复制这种巨大的RNA（大约29种内切酶）。其中许多在复制（和/或转录）中的作用是未知的。我们试图从它们的三维结构中得出有关这些蛋白质可能功能的结论，我们通过X射线晶体学确定它们的三维结构。非结构蛋白3在其1922个氨基酸的多肽链中含有至少7个不同的功能模块。其中之一是所谓的SARS独特结构域，这是一段约338个残基的片段，在任何其他冠状病毒中都完全不存在。因此，与该家族的其他病毒相比，SARS冠状病毒的致病性非常高，这可能是其原因。我们在这里描述的三维结构的SARS独特的域，并表明，它由两个模块与一个已知的倍，所谓的宏域。此外，我们证明，这些结构域结合不寻常的核酸结构形成的连续的鸟苷核苷酸，其中四条链的核酸形成超螺旋（所谓的G-四链体）。SUD可能参与与具有这种特殊结构的病毒或宿主细胞RNA的结合，从而调节病毒复制或对抗受感染宿主细胞的免疫反应。
Since the outbreak of severe acute respiratory syndrome (SARS) in 2003, the three-dimensional structures of several of the replicase/transcriptase components of SARS coronavirus (SARS-CoV), the non-structural proteins (Nsps), have been determined. However, within the large Nsp3 (1922 amino-acid residues), the structure and function of the so-called SARS-unique domain (SUD) have remained elusive. SUD occurs only in SARS-CoV and the highly related viruses found in certain bats, but is absent from all other coronaviruses. Therefore, it has been speculated that it may be involved in the extreme pathogenicity of SARS-CoV, compared to other coronaviruses, most of which cause only mild infections in humans. In order to help elucidate the function of the SUD, we have determined crystal structures of fragment 389–652 (“SUDcore”) of Nsp3, which comprises 264 of the 338 residues of the domain. Both the monoclinic and triclinic crystal forms (2.2 and 2.8 Å resolution, respectively) revealed that SUDcore forms a homodimer. Each monomer consists of two subdomains, SUD-N and SUD-M, with a macrodomain fold similar to the SARS-CoV X-domain. However, in contrast to the latter, SUD fails to bind ADP-ribose, as determined by zone-interference gel electrophoresis. Instead, the entire SUDcore as well as its individual subdomains interact with oligonucleotides known to form G-quadruplexes. This includes oligodeoxy- as well as oligoribonucleotides. Mutations of selected lysine residues on the surface of the SUD-N subdomain lead to reduction of G-quadruplex binding, whereas mutations in the SUD-M subdomain abolish it. As there is no evidence for Nsp3 entering the nucleus of the host cell, the SARS-CoV genomic RNA or host-cell mRNA containing long G-stretches may be targets of SUD. The SARS-CoV genome is devoid of G-stretches longer than 5–6 nucleotides, but more extended G-stretches are found in the 3′-nontranslated regions of mRNAs coding for certain host-cell proteins involved in apoptosis or signal transduction, and have been shown to bind to SUD in vitro. Therefore, SUD may be involved in controlling the host cell's response to the viral infection. Possible interference with poly(ADP-ribose) polymerase-like domains is also discussed. The genome of the SARS coronavirus codes for 16 non-structural proteins that are involved in replicating this huge RNA (approximately 29 kilobases). The roles of many of these in replication (and/or transcription) are unknown. We attempt to derive conclusions concerning the possible functions of these proteins from their three-dimensional structures, which we determine by X-ray crystallography. Non-structural protein 3 contains at least seven different functional modules within its 1922-amino-acid polypeptide chain. One of these is the so-called SARS-unique domain, a stretch of about 338 residues that is completely absent from any other coronavirus. It may thus be responsible for the extraordinarily high pathogenicity of the SARS coronavirus, compared to other viruses of this family. We describe here the three-dimensional structure of the SARS-unique domain and show that it consists of two modules with a known fold, the so-called macrodomain. Furthermore, we demonstrate that these domains bind unusual nucleic-acid structures formed by consecutive guanosine nucleotides, where four strands of nucleic acid are forming a superhelix (so-called G-quadruplexes). SUD may be involved in binding to viral or host-cell RNA bearing this peculiar structure and thereby regulate viral replication or fight the immune response of the infected host cell.
登录
查看更多内容
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
56.9
作者：
Anand, K;Ziebuhr, J;Hilgenfeld, R
通讯作者：
Hilgenfeld, R
DOI：
10.1073/pnas.241374798
发表时间：
2001-11-20
影响因子：
11.1
作者：
Deng, JP;Xiong, Y;Sundaralingam, M
通讯作者：
Sundaralingam, M
影响因子：
64.5
作者：
Imai Y;Kuba K;Neely GG;Yaghubian-Malhami R;Perkmann T;van Loo G;Ermolaeva M;Veldhuizen R;Leung YH;Wang H;Liu H;Sun Y;Pasparakis M;Kopf M;Mech C;Bavari S;Peiris JS;Slutsky AS;Akira S;Hultqvist M;Holmdahl R;Nicholls J;Jiang C;Binder CJ;Penninger JM
通讯作者：
Penninger JM
影响因子：
2.5
作者：
Hakme, Antoinette;Huber, Aline;Schreiber, Valerie
通讯作者：
Schreiber, Valerie