CryoEM and AI reveal a structure of SARS-CoV-2 Nsp2, a multifunctional protein involved in key host processes
This article has been Reviewed by the following groups
Listed in
- Evaluated articles (ScreenIT)
Abstract
The SARS-CoV-2 protein Nsp2 has been implicated in a wide range of viral processes, but its exact functions, and the structural basis of those functions, remain unknown. Here, we report an atomic model for full-length Nsp2 obtained by combining cryo-electron microscopy with deep learning-based structure prediction from AlphaFold2. The resulting structure reveals a highly-conserved zinc ion-binding site, suggesting a role for Nsp2 in RNA binding. Mapping emerging mutations from variants of SARS-CoV-2 on the resulting structure shows potential host-Nsp2 interaction regions. Using structural analysis together with affinity tagged purification mass spectrometry experiments, we identify Nsp2 mutants that are unable to interact with the actin-nucleation-promoting WASH protein complex or with GIGYF2, an inhibitor of translation initiation and modulator of ribosome-associated quality control. Our work suggests a potential role of Nsp2 in linking viral transcription within the viral replication-transcription complexes (RTC) to the translation initiation of the viral message. Collectively, the structure reported here, combined with mutant interaction mapping, provides a foundation for functional studies of this evolutionary conserved coronavirus protein and may assist future drug design.
Article activity feed
-
SciScore for 10.1101/2021.05.10.443524: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
NIH rigor criteria are not applicable to paper type.Table 2: Resources
Recombinant DNA Sentences Resources Nsp2 Expression: SARS-CoV-2 Nsp2, codon optimized for Escherichia coli expression, was cloned in a pET-29b(+) vector backbone with N-terminus 10XHis-tag and SUMO-tag (Ep156). pET-29b(+)suggested: NoneSoftware and Algorithms Sentences Resources Each collection was performed with semi-automated scripts in SerialEM. SerialEMsuggested: (SerialEM, RRID:SCR_017293)Cys/His residues were manually identified for Zn coordination sites and Zn was placed in COOT. COOTsuggested: (Coot, RRID:SCR_014222)Local resolution was determined by running ResMap program45. ResMapsuggested: NoneFirst three were individually … SciScore for 10.1101/2021.05.10.443524: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
NIH rigor criteria are not applicable to paper type.Table 2: Resources
Recombinant DNA Sentences Resources Nsp2 Expression: SARS-CoV-2 Nsp2, codon optimized for Escherichia coli expression, was cloned in a pET-29b(+) vector backbone with N-terminus 10XHis-tag and SUMO-tag (Ep156). pET-29b(+)suggested: NoneSoftware and Algorithms Sentences Resources Each collection was performed with semi-automated scripts in SerialEM. SerialEMsuggested: (SerialEM, RRID:SCR_017293)Cys/His residues were manually identified for Zn coordination sites and Zn was placed in COOT. COOTsuggested: (Coot, RRID:SCR_014222)Local resolution was determined by running ResMap program45. ResMapsuggested: NoneFirst three were individually aligned with matchmaker in ChimeraX to the experimental model to assess the similarity and report RMSDs in the main text. ChimeraXsuggested: (UCSF ChimeraX, RRID:SCR_015872)Sequence alignment and sequence conservation analysis: Nsp2 sequences were manually downloaded from UniProt and aligned in Jalview using MAFFT49,50. UniProtsuggested: (UniProtKB, RRID:SCR_004426)Jalviewsuggested: (Jalview, RRID:SCR_006459)Conservation was mapped on the Nsp2 structure using a combination of Chimera and ChimeraX and the supplementary alignment figure was prepared with MView server47,51,52. Conservationsuggested: (Conservation, RRID:SCR_016064)Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
Results from scite Reference Check: We found no unreliable references.
-
