Proteopedia

SARS-CoV-2 enzyme 2'-O-MT

Non-structural protein 16 (Nsp16): 2'-O-methyltransferase (2'-O-MT)

Nsp 16 is a methyltransferase enzyme that mediates mRNA cap 2'-O-ribose methylation to the 5'-cap structure of viral mRNAs. The N7-methyl guanosine cap is a prerequisite for binding of nsp16, therefore plays an essential role in viral mRNAs cap methylation which is essential to evade the host immune system.[1][2]

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FunctionFunction

SARS-CoV-2 non-structural protein 16 (nsp16) is a ribose 2’-O-methyltransferase that forms a heterodimer together with its allosteric activator nsp10[3]. Nsp16 is created as part of the polyprotein pp1ab and consists of 298 residues[4]. The role nsp16 plays in the virus’s life cycle is to perform the final step of RNA cap synthesis. Capping of the 5’-end of mRNA stabilizes it, preventing degradation by the host cell and helps to reduce an innate immune response[3]. Further, this 2’-O-methylation is important for effective RNA translation[5]. The nsp10-nsp16 complex modifies the mRNA’s cap-0 structure, which was previously methylated by the nsp14 (SARS-CoV-2_enzyme_ExoN), another S-adenosyl methionine (SAM)-dependent methyltransferase[3]. Nsp16 then converts the cap-0 (m7GpppN-RNA) to a cap-1 structure (m7GpppNm-RNA) by adding a methyl group at the ribose 2’-O position of the first nucleotide using SAM as a methyl donor[3][5]. Nsp10 works as a co-factor for nsp16, stabilizing the SAM-binding pocket[4] and enhancing the enzymatic activity significantly[5]. For SARS-CoV, and similarly for MERS-CoV, the affinity for m7GpppA-RNA and m7GpppA cap analogue of nsp16 was found to be low until binding to nsp10, which enhances the affinity for binding to RNA[3] [6].

DiseaseDisease

SARS-CoV-2 is cause of the global COVID-19 pandemic.

StructureStructure

The secondary structure of nsp16 is composed of 12 β-strands and 12 α-helices[5]. Its center is comprised of a twisted β-sheet, consisting of 8 β-strands (β4, β3, β2, β6, β7, β9, β8, β1), where β9 and β1 are antiparallel to the other 6[7]. This β-sheet is surrounded by α-helices α5-α9, with helices α3 and α4 stabilizing it from the nsp10-nsp16 interface. This interface consists of β4, α3, α4 and α10 of nsp16 and α2, α3 and α4 of nsp10, and takes up an area of 1983 Å^2[5]. Nsp16 has a central canyon enriched with negatively charged residues made up of the loops of the C-terminal ends of β2, β3, β4 and β6 that form a SAM binding pocket[7]. The RNA binding site is a positively charged groove near the SAM binding pocket[5]. The amino acids 20-40 and 133-143 work as gate loops 1 and 2, rotating when the site binds to a cap-0 structure and resulting in a widening of the RNA binding pocket. These loops form a groove to accommodate the substrate[7]. All current structures of nsp16 are in complex with nsp10 and the best resolved structure from SARS-CoV-2 to date is 6w4h with 1.80 Å resolution.

VariationsVariations

The sequences of nsp10 and nsp16 are both highly conserved within the beta coronaviruses with nsp16 having 95% amino acid sequence identity between SARS-CoV and SARS-CoV-2[4]. Notable highly conserved regions are the KDKE motif, a catalytic tetrad consisting of Lys46, Asp130, Lys170 and Glu203, as well as the residues responsible for the substrate binding of SAM[3] [4]. Most residue variations between strains are not found in the catalytic or substrate binding sites or the nsp10 interface, but instead at the surface of the protein exposed to the solvent[4]. An exception are some mutations in the sequence of gate loops 1 and 2 compared to SARS-CoV-1, which might have an impact on RNA binding or kinetics of the enzyme. Besides other mutations nsp16 might have the ability to bind small molecules with a heterocyclic ring in a adenosine binding pocket ~25 Å distant from the catalytic pocket compared to nsp16 of SARS-CoV[7].

See alsoSee also

Coronavirus_Disease 2019 (COVID-19)

ReferencesReferences

  1. ↑ Modeling of the SARS-COV-2 Genome
  2. ↑ Zhang C, Zheng W, Huang X, Bell EW, Zhou X, Zhang Y. Protein Structure and Sequence Reanalysis of 2019-nCoV Genome Refutes Snakes as Its Intermediate Host and the Unique Similarity between Its Spike Protein Insertions and HIV-1. J Proteome Res. 2020 Apr 3;19(4):1351-1360. doi: 10.1021/acs.jproteome.0c00129., Epub 2020 Mar 24. PMID:32200634 doi:http://dx.doi.org/10.1021/acs.jproteome.0c00129
  3. ↑ 3.0 3.1 3.2 3.3 3.4 3.5 Romano M, Ruggiero A, Squeglia F, Maga G, Berisio R. A Structural View of SARS-CoV-2 RNA Replication Machinery: RNA Synthesis, Proofreading and Final Capping. Cells. 2020 May 20;9(5). pii: cells9051267. doi: 10.3390/cells9051267. PMID:32443810 doi:http://dx.doi.org/10.3390/cells9051267
  4. ↑ 4.0 4.1 4.2 4.3 4.4 Rosas-Lemus M, Minasov G, Shuvalova L, Inniss NL, Kiryukhina O, Wiersum G, Kim Y, Jedrzejczak R, Maltseva NI, Endres M, Jaroszewski L, Godzik A, Joachimiak A, Satchell KJF. The crystal structure of nsp10-nsp16 heterodimer from SARS-CoV-2 in complex with S-adenosylmethionine. bioRxiv. 2020 Apr 26. doi: 10.1101/2020.04.17.047498. PMID:32511376 doi:http://dx.doi.org/10.1101/2020.04.17.047498
  5. ↑ 5.0 5.1 5.2 5.3 5.4 5.5 Krafcikova P, Silhan J, Nencka R, Boura E. Structural analysis of the SARS-CoV-2 methyltransferase complex involved in RNA cap creation bound to sinefungin. Nat Commun. 2020 Jul 24;11(1):3717. doi: 10.1038/s41467-020-17495-9. PMID:32709887 doi:http://dx.doi.org/10.1038/s41467-020-17495-9
  6. ↑ Chen Y, Su C, Ke M, Jin X, Xu L, Zhang Z, Wu A, Sun Y, Yang Z, Tien P, Ahola T, Liang Y, Liu X, Guo D. Biochemical and structural insights into the mechanisms of SARS coronavirus RNA ribose 2'-O-methylation by nsp16/nsp10 protein complex. PLoS Pathog. 2011 Oct;7(10):e1002294. doi: 10.1371/journal.ppat.1002294. Epub, 2011 Oct 13. PMID:22022266 doi:http://dx.doi.org/10.1371/journal.ppat.1002294
  7. ↑ 7.0 7.1 7.2 7.3 Viswanathan T, Arya S, Chan SH, Qi S, Dai N, Misra A, Park JG, Oladunni F, Kovalskyy D, Hromas RA, Martinez-Sobrido L, Gupta YK. Structural basis of RNA cap modification by SARS-CoV-2. Nat Commun. 2020 Jul 24;11(1):3718. doi: 10.1038/s41467-020-17496-8. PMID:32709886 doi:http://dx.doi.org/10.1038/s41467-020-17496-8

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