High-resolution structures of the SARS-CoV-2 N7-methyltransferase inform therapeutic development

Kottur, J.; Rechkoblit, Olga; Quintana-Feliciano, Richard; Sciaky, Daniela; Aggarwal, Aneel K.

doi:10.1038/s41594-022-00828-1

Cited by 16 publications

(24 citation statements)

References 25 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Very recently, several crystal structures of full-length nsp14 from SARS-CoV-2 were solved, bound to SAH (PDB 7R2V , 7QGI ) or to a cap-analogue (PDB 7QIF ) [ 27 , 28 ]. A high-resolution crystal structure of the SARS-CoV-2 N 7-MTase domain - expressed in fusion with the TELSAM epitope - was also reported [ 29 ]. These new structures provide crucial information for understanding the dynamics of nsp14 methyltransferase activity and pave the way for the structure-based development of new antivirals against CoVs.…”

Section: Introductionmentioning

confidence: 99%

Structure-guided optimization of adenosine mimetics as selective and potent inhibitors of coronavirus nsp14 N7-methyltransferases

Hausdorff,

Delpal,

Barelier

et al. 2023

European Journal of Medicinal Chemistry

View full text Add to dashboard Cite

Section: Introductionmentioning

confidence: 99%

Structure-guided optimization of adenosine mimetics as selective and potent inhibitors of coronavirus nsp14 N7-methyltransferases

Hausdorff,

Delpal,

Barelier

et al. 2023

European Journal of Medicinal Chemistry

View full text Add to dashboard Cite

“…Data-processing and refinement statistics for this structure are given in Table 2. This observation suggests that fusion to 1TEL improved the resolution of vWa crystals produced in our hands by an average of 0.4 A ˚, a significant improvement (Table 1), providing an exciting hint into what more experienced crystallographers might be able to accomplish with TELSAM-mediated crystallization, such as in a recent stunning example involving the SARS-CoV2 nsp14 N7-methyltransferase domain (Kottur et al, 2022).…”

Section: A Bulkier Telsam-target Linker Did Not Alter the Crystalpack...mentioning

confidence: 63%

“…This parallels the experience of Kottur and coworkers, where three different linkers were tested in an attempt to crystallize the recalcitrant SARS CoV2 nsp14 N7-methyltransferase. In this case, only one of the three linkers resulted in crystals, but with optimization of the crystallization conditions the authors were able to reliably achieve a diffraction resolution of 1.4-1.6 A ˚ (Kottur et al, 2022).…”

Section: Discussionmentioning

confidence: 99%

“…Fusion to TELSAM has now been demonstrated in multiple cases to allow the rapid crystallization of even difficult target proteins (Kottur et al, 2022), to enable high-resolution data collection (<2.0 A ˚), to allow crystallization at unusually low protein concentrations and to result in crystals where the protein of interest has an increased degree of residual molecular motion, as shown by significantly higher crystallographic B factors. These higher B factors hint that TELSAM-mediated crystals may capture proteins of interest in subtly more physiological conformations, with less interference from crystal-packing artefacts.…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Increasing the bulk of the 1TEL–target linker and retaining the 10×His tag in a 1TEL–CMG2-vWa construct improves crystal order and diffraction limits

Gajjar,

Pedroza Romo,

Litchfield

et al. 2023

Acta Cryst Sect D Struct Biol

View full text Add to dashboard Cite

TELSAM-fusion crystallization has the potential to become a revolutionary tool for the facile crystallization of proteins. TELSAM fusion can increase the crystallization rate and enable crystallization at low protein concentrations, in some cases with minimal crystal contacts [Nawarathnage et al. (2022), Open Biol. 12, 210271]. Here, requirements for the linker composition between 1TEL and a fused CMG2 vWa domain were investigated. Ala-Ala, Ala-Val, Thr-Val and Thr-Thr linkers were evaluated, comparing metrics for crystallization propensity and crystal order. The effect on crystallization of removing or retaining the purification tag was then tested. It was discovered that increasing the linker bulk and retaining the 10×His purification tag improved the diffraction resolution, likely by decreasing the number of possible vWa-domain orientations in the crystal. Additionally, it was discovered that some vWa-domain binding modes are correlated with scrambling of the 1TEL polymer orientation in crystals and an effective mitigation strategy for this pathology is presented.

show abstract

“…The SARS-CoV-2 nsp14 MTase domain-encoding sequence (GenBank: YP 009725309.1, residues 300-527) was codon-optimized for expression in E. coli and synthesized by GeneArt (Thermo Scientific). The gene was cloned into a modified pRSFDuet vector containing an N-terminal hexahistidine (His6) purification tag, followed by a SUMO solubility and folding tag, and a TELSAM crystallization tag [38]. The plasmid was transformed into E. coli BL21 DE3 NiCo bacterial strain (New England Biolabs), and the protein was overexpressed using autoinduction ZY medium.…”

Section: Protein Expression and Purificationmentioning

confidence: 99%

Drug Discovery in Low Data Regimes: Leveraging a Computational Pipeline for the Discovery of Novel SARS-CoV-2 Nsp14-MTase Inhibitors

Nigam,

Hurley,

et al. 2023

Preprint

View full text Add to dashboard Cite

The COVID-19 pandemic, caused by the SARS-CoV-2 virus, has led to significant global morbidity and mortality. A crucial viral protein, the non-structural protein 14 (nsp14), catalyzes the methylation of viral RNA and plays a critical role in viral genome replication and transcription. Due to the low mutation rate in the nsp region among various SARS-CoV-2 variants, nsp14 has emerged as a promising therapeutic target. However, discovering potential inhibitors remains a challenge. In this work, we introduce a computational pipeline for the rapid and efficient identification of potential nsp14 inhibitors by leveraging virtual screening and the NCI open compound collection, which contains 250,000 freely available molecules for researchers worldwide. The introduced pipeline provides a cost-effective and efficient approach for early-stage drug discovery by allowing researchers to evaluate promising molecules without incurring synthesis expenses. Our pipeline successfully identified seven promising candidates after experimentally validating only 40 compounds. Notably, we discovered NSC620333, a compound that exhibits a strong binding affinity to nsp14 with a dissociation constant of 427 ± 84 nM. In addition, we gained new insights into the structure and function of this protein through molecular dynamics simulations. We identified new conformational states of the protein and determined that residues Phe367, Tyr368, and Gln354 within the binding pocket serve as stabilizing residues for novel ligand interactions. We also found that metal coordination complexes are crucial for the overall function of the binding pocket. Lastly, we present the solved crystal structure of the nsp14-MTase complexed with SS148 (PDB:8BWU), a potent inhibitor of methyltransferase activity at the nanomolar level (IC50value of 70 ± 6 nM). Our computational pipeline accurately predicted the binding pose of SS148, demonstrating its effectiveness and potential in accelerating drug discovery efforts against SARS-CoV-2 and other emerging viruses.

show abstract

High-resolution structures of the SARS-CoV-2 N7-methyltransferase inform therapeutic development

Cited by 16 publications

References 25 publications

Structure-guided optimization of adenosine mimetics as selective and potent inhibitors of coronavirus nsp14 N7-methyltransferases

Structure-guided optimization of adenosine mimetics as selective and potent inhibitors of coronavirus nsp14 N7-methyltransferases

Increasing the bulk of the 1TEL–target linker and retaining the 10×His tag in a 1TEL–CMG2-vWa construct improves crystal order and diffraction limits

Drug Discovery in Low Data Regimes: Leveraging a Computational Pipeline for the Discovery of Novel SARS-CoV-2 Nsp14-MTase Inhibitors

Contact Info

Product

Resources

About