The SARS-Unique Domain (SUD) of SARS Coronavirus Contains Two Macrodomains That Bind G-Quadruplexes

Abstract

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 SARSunique 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 (''SUD core '') 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 SUD core 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 SUD core 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 Gquadruplex 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 39-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.

Structural and biophysical analysis of sequence insertions in the Venezuelan Equine Encephalitis Virus macro domain

“…To date, the role of viral macro domains has been demonstrated in the modification of cellular response to viral infection (Nan et al, 2014;Nargi-Aizenman et al, 2002;Tan et al, 2009), as a possible RNA recruitment factor (Malet et al, 2009), or in minus-strand synthesis, P23 processing and/or viral replication (LaStarza et al, 1994a(LaStarza et al, , 1994bLulla et al, 2011). The molecular mechanism or biochemical function behind this role is not well understood.…”

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“…Coronaviridae that do not present it (65). Additionally, it has been suggested that the SUD domain interacts with G-quadruplexes of the host.…”

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“…Macrodomains are found in proteins from all kingdoms of life supporting different cellular processes . Macrodomain‐containing proteins can recognize variety of substrates, including MARylated and PARylated proteins, different ADPr metabolites (such as O AADPr) and RNA (Fig. ).…”