Structural basis for RNA-genome recognition during bacteriophage Qβ replication

Gytz, Heidi; Mohr, Durita; Seweryn, Paulina; Yoshimura, Yuichi; Kutlubaeva, Zarina; Dolman, Fleur; Chelchessa, Bosene; Chetverin, Alexander B.; Mulder, Frans A. A.; Brodersen, D.E.; Knudsen, Charlotte R.

doi:10.1093/nar/gkv1212

Cited by 23 publications

(28 citation statements)

References 51 publications

(90 reference statements)

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“…During infection in bacteria, Qb recruits the host-cell ribosomal S1 protein, which is composed of six oligonucleotide-oligosaccharide-binding (OB) domains, to initiate viral replication (Wahba et al, 1974). Two of these domains, OB1 and OB2, contact the Qb replicase core but use different motifs to contact discrete regions and wrap around the replicase core (Gytz et al, 2015;Takeshita et al, 2014). Thus, although the structure of S1 and P share no homology, the modular domains of S1 and three of the monomers of P contact similar regions on the finger subdomain of their respective RdRp using a similar strategy ( Figure S7).…”

Section: Discussionmentioning

confidence: 99%

Structure of the Respiratory Syncytial Virus Polymerase Complex

Gilman

Liu

Fung

et al. 2019

Cell

134

209

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Graphical AbstractHighlights d Cryo-EM structure of RSV L bound by tetrameric RSV P solved to 3.2 Å d P tetramer adopts an asymmetric tentacular arrangement when bound to L d L priming loop adopts elongation-compatible state without PRNTase-RdRp separation d Structure rationalizes escape from small-molecule antivirals SUMMARY Numerous interventions are in clinical development for respiratory syncytial virus (RSV) infection, including small molecules that target viral transcription and replication. These processes are catalyzed by a complex comprising the RNA-dependent RNA polymerase (L) and the tetrameric phosphoprotein (P). RSV P recruits multiple proteins to the polymerase complex and, with the exception of its oligomerization domain, is thought to be intrinsically disordered. Despite their critical roles in RSV transcription and replication, structures of L and P have remained elusive. Here, we describe the 3.2-Å cryo-EM structure of RSV L bound to tetrameric P. The structure reveals a striking tentacular arrangement of P, with each of the four monomers adopting a distinct conformation. The structure also rationalizes inhibitor escape mutants and mutations observed in live-attenuated vaccine candidates. These results provide a framework for determining the molecular underpinnings of RSV replication and transcription and should facilitate the design of effective RSV inhibitors.

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Section: Discussionmentioning

confidence: 99%

Structure of the Respiratory Syncytial Virus Polymerase Complex

Gilman

Liu

Fung

et al. 2019

Cell

134

209

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“…single-particle cryo-EM | ssRNA virus | genome packaging | maturation protein | lysis protein S ingle-stranded RNA (ssRNA) viruses infect all domains of life and are important pathogens (1)(2)(3)(4). Bacteriophages or phages, especially the canonical Leviviridae Qβ and MS2, have been model systems for studying ssRNA viral gene regulation (5)(6)(7)(8), genome replication (9,10), and encapsidation (11)(12)(13)(14). They have a near-icosahedral capsid, encapsidating genomic RNAs (gRNAs) of ∼3,500-4,300 nucleotides.…”

mentioning

confidence: 99%

Structures of Qβ virions, virus-like particles, and the Qβ–MurA complex reveal internal coat proteins and the mechanism of host lysis

Zhao

Gorzelnik

Chang

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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In single-stranded RNA bacteriophages (ssRNA phages) a single copy of the maturation protein binds the genomic RNA (gRNA) and is required for attachment of the phage to the host pilus. For the canonical Qβ the maturation protein, A, has an additional role as the lysis protein, by its ability to bind and inhibit MurA, which is involved in peptidoglycan biosynthesis. Here, we determined structures of Qβ virions, virus-like particles, and the Qβ-MurA complex using single-particle cryoelectron microscopy, at 4.7-Å, 3.3-Å, and 6.1-Å resolutions, respectively. We identified the outer surface of the β-region in A as the MurA-binding interface. Moreover, the pattern of MurA mutations that block Qβ lysis and the conformational changes of MurA that facilitate A binding were found to be due to the intimate fit between A and the region encompassing the closed catalytic cleft of substrate-liganded MurA. Additionally, by comparing the Qβ virion with Qβ virus-like particles that lack a maturation protein, we observed a structural rearrangement in the capsid coat proteins that is required to package the viral gRNA in its dominant conformation. Unexpectedly, we found a coat protein dimer sequestered in the interior of the virion. This coat protein dimer binds to the gRNA and interacts with the buried α-region of A, suggesting that it is sequestered during the early stage of capsid formation to promote the gRNA condensation required for genome packaging. These internalized coat proteins are the most asymmetrically arranged major capsid proteins yet observed in virus structures.

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“…An analysis of the amino acid sequence [ 6 ] has led to the conclusion that there are six OB domains in protein S1 ( Figure ). Subsequent structural studies have confirmed this conclusion with further refinement: the N-terminal OB domain (OB 1 ) contains 4 rather than 5 β-strands [ 10 , 16 - 18 ].…”

Section: Domain Structure Of Protein S1mentioning

confidence: 98%

“…At the same time, it was thought that the domains OB1 and OB2 do not bind RNA and are involved in the protein-protein interactions responsible for the binding of protein S1 to the ribosome and to the Qβ replicase core [ 1 ]. Recent studies have demonstrated that the OB1 and OB2 domains indeed form contacts with the Qβ replicase core [ 17 , 18 ], but, in addition, domain OB2 can interact with RNA due to the high density of positively charged residues on the surface area not involved in the protein-protein interaction [ 18 ]. Thus, all five classical OB domains of S1 possess RNA-binding properties.…”

Section: Domain Structure Of Protein S1mentioning

confidence: 99%

The Contribution of Ribosomal Protein S1 to the Structure and Function of Qβ Replicase

Kutlubaeva¹,

Chetverina²,

Chetverin³

2017

Acta Naturae

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The high resolution crystal structure of bacterial ribosome was determined more than 10 years ago; however, it contains no information on the structure of the largest ribosomal protein, S1. This unusual protein comprises six flexibly linked domains; therefore, it lacks a fixed structure and this prevents the formation of crystals. Besides being a component of the ribosome, protein S1 also serves as one of the four subunits of Qβ replicase, the RNA-directed RNA polymerase of bacteriophage Qβ. In each case, the role of this RNA-binding protein has been thought to consist in holding the template close to the active site of the enzyme. In recent years, a breakthrough was made in studies of protein S1 within Qβ replicase. This includes the discovery of its paradoxical ability to displace RNA from the replicase complex and determining the crystal structure of its fragment capable of performing this function. The new findings call for a re-examination of the contribution of protein S1 to the structure and function of the ribosome.

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Structural basis for RNA-genome recognition during bacteriophage Qβ replication

Cited by 23 publications

References 51 publications

Structure of the Respiratory Syncytial Virus Polymerase Complex

Structure of the Respiratory Syncytial Virus Polymerase Complex

Structures of Qβ virions, virus-like particles, and the Qβ–MurA complex reveal internal coat proteins and the mechanism of host lysis

The Contribution of Ribosomal Protein S1 to the Structure and Function of Qβ Replicase

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