3C-like protease

{{Short description|Class of enzymes}} {{see also|COVID-19 drug development}} {{Infobox enzyme | Name = SARS coronavirus main proteinase | EC_number = 3.4.22.69 | CAS_number = | GO_code = | image = File:DOI.10.1126.science.abb4489.S2.png | width = 258 | caption = SARS-CoV-2 main proteinase dimer with the catalytic dyad (H41; C145) in complex with a covalent peptidomimetic protease inhibitor ("11a", magenta). From {{PDB|6LZE}}.<ref name="pmid32321856" /> | name = }} {{Infobox protein family |Name=Peptidase C30, Coronavirus endopeptidase |InterPro=IPR008740 |Pfam=PF05409 |PROSITE=PS51442 |Symbol=Peptidase_C30 |MEROPS=C30 |SCOP=d1q2wb1 }} The '''3C-like protease''' ('''3CLpro''') or '''main protease''' ('''Mpro'''), formally known as '''C30 endopeptidase''',<ref>{{cite journal |vauthors=Ahmad B, Batool M, Ain QU, Kim MS, Choi S |date=August 2021 |title=Exploring the Binding Mechanism of PF-07321332 SARS-CoV-2 Protease Inhibitor through Molecular Dynamics and Binding Free Energy Simulations |journal=International Journal of Molecular Sciences |volume=22 |issue=17 |page=9124 |doi=10.3390/ijms22179124 |pmc=8430524 |pmid=34502033 |doi-access=free}}</ref> is the main protease found in coronaviruses. It cleaves the coronavirus polyprotein at eleven conserved sites. It is a cysteine protease and a member of the PA clan of proteases. It has a cysteine-histidine catalytic dyad at its active site and cleaves a Gln–(Ser/Ala/Gly) peptide bond.

The Enzyme Commission refers to this family as '''SARS coronavirus main proteinase''' ('''Mpro'''; {{EC number|3.4.22.69}}). The 3CL protease corresponds to coronavirus '''nonstructural protein 5''' (nsp5). The "3C" in the common name refers to the 3C protease (3Cpro) which is a homologous protease found in picornaviruses.

== Function ==

The 3C-like protease is able to catalytically cleave a peptide bond between a glutamine at position P1 and a small amino acid (serine, alanine, or glycine) at position P1'. The SARS coronavirus 3CLpro can for instance self-cleave the following peptides:<ref>{{cite journal |vauthors =Goetz DH, Choe Y, Hansell E, Chen YT, McDowell M, Jonsson CB, Roush WR, McKerrow J, Craik CS |title =Substrate specificity profiling and identification of a new class of inhibitor for the major protease of the SARS coronavirus |journal =Biochemistry |volume =46 |issue =30 |pages =8744–52 |date =July 2007 |pmid =17605471 |doi =10.1021/bi0621415 }}</ref><ref name=":0">{{cite journal |vauthors =Fan K, Wei P, Feng Q, Chen S, Huang C, Ma L, Lai B, Pei J, Liu Y, Chen J, Lai L |title =Biosynthesis, purification, and substrate specificity of severe acute respiratory syndrome coronavirus 3C-like proteinase |journal =The Journal of Biological Chemistry |volume =279 |issue =3 |pages =1637–42 |date =January 2004 |pmid =14561748 |doi =10.1074/jbc.m310875200 |pmc =7980035 |doi-access =free }}</ref><ref>{{cite journal |vauthors =Akaji K, Konno H, Onozuka M, Makino A, Saito H, Nosaka K |title =Evaluation of peptide-aldehyde inhibitors using R188I mutant of SARS 3CL protease as a proteolysis-resistant mutant |journal =Bioorganic & Medicinal Chemistry |volume =16 |issue =21 |pages =9400–8 |date =November 2008 |pmid =18845442 |doi =10.1016/j.bmc.2008.09.057 |pmc =7126698 }}</ref>

{{blockquote|TSAVLQ-SGFRK-NH2 and SGVTFQ-GKFKK are the two peptides corresponding to the two self-cleavage sites of the SARS 3C-like proteinase|author=|title=|source=}}

The protease is important in the processing of the coronavirus replicase polyprotein ({{UniProt|P0C6U8}}). It is the main protease in coronaviruses and corresponds to nonstructural protein 5 (nsp5).<ref name="Fehr_2015">{{cite book |title=Coronaviruses |vauthors=Fehr AR, Perlman S |date=2015 |publisher=Springer |isbn=978-1-4939-2438-7 |veditors=Maier HJ, Bickerton E, Britton P |series=Methods in Molecular Biology |volume=1282 |pages=1–23 |chapter=Coronaviruses: an overview of their replication and pathogenesis |doi=10.1007/978-1-4939-2438-7_1 |pmc=4369385 |pmid=25720466 |quote=See section: Virion Structure.}}</ref> It cleaves the coronavirus polyprotein at 11 conserved sites. The 3CL protease has a cysteine-histidine catalytic dyad at its active site.<ref name=":0" /> The sulfur of the cysteine acts as a nucleophile and the imidazole ring of the histidine as a general base.<ref name="pmid20167482">{{cite journal |display-authors=6 |issn=0960-894X |vauthors =Ryu YB, Park SJ, Kim YM, Lee JY, Seo WD, Chang JS, Park KH, Rho MC, Lee WS |title =SARS-CoV 3CLpro inhibitory effects of quinone-methide triterpenes from Tripterygium regelii |journal =Bioorganic & Medicinal Chemistry Letters |volume =20 |issue =6 |pages =1873–6 |date =March 2010 |pmid =20167482 |doi =10.1016/j.bmcl.2010.01.152 |pmc =7127101 }}</ref> {| class="wikitable" |+ Substrate preferences for 3CL proteases (from table 2)<ref name="pmid22073294">{{cite journal |vauthors =Chuck CP, Chow HF, Wan DC, Wong KB |title =Profiling of substrate specificities of 3C-like proteases from group 1, 2a, 2b, and 3 coronaviruses |journal =PLOS ONE |volume =6 |issue =11 |article-number =e27228 |date =2011 |pmid =22073294 |doi =10.1371/journal.pone.0027228 |pmc =3206940 |doi-access =free |bibcode =2011PLoSO...627228C }}</ref> |- ! Position !! Substrate preference |- | P5 || No strong preference |- | P4 || Small hydrophobic residues |- | P3 || Positively charged residue |- | P2 || High hydrophobicity and absence of beta-branch |- | P1 || Glutamine |- | P1' || Small residues |- | P2' || Small residues |- | P3' || No strong preference |}

== Nomenclature ==

Alternative names provided by the EC include ''3CLpro'', ''3C-like protease'', ''coronavirus 3C-like protease'', ''Mpro'', ''SARS 3C-like protease'', ''SARS coronavirus 3CL protease'', ''SARS coronavirus main peptidase'', ''SARS coronavirus main protease'', ''SARS-CoV 3CLpro enzyme'', ''SARS-CoV main protease'', ''SARS-CoV Mpro'' and ''severe acute respiratory syndrome coronavirus main protease''.

== As a treatment target == [[Image:Nirtalmatrelvir on 3CL.png|thumb|left|Nirmatrelvir bound to 3CL {{PDB|7RFW}}]] [[Image:PF-07321332.svg|thumb|left|Nirmatrelvir, a 3CLpro inhibitor developed by Pfizer in phase II/III clinical trials as a combination drug with ritonavir.<ref name="pmid34029993">{{cite journal |vauthors =Vandyck K, Deval J |title =Considerations for the discovery and development of 3-chymotrypsin-like cysteine protease inhibitors targeting SARS-CoV-2 infection |journal =Curr Opin Virol |volume =49 |pages =36–40 |date =August 2021 |pmid =34029993 |doi =10.1016/j.coviro.2021.04.006 |pmc =8075814 }}</ref><ref name="arena">{{cite web |url =https://www.clinicaltrialsarena.com/news/pfizer-antiviral-covid-trial/ |title =Pfizer begins dosing in Phase II/III trial of antiviral drug for Covid-19. |work =Clinical Trials Arena |date =2 September 2021 }}</ref>]] A number of protease inhibitors are being developed targeting 3CLpro and homologous 3Cpro, including CLpro-1, GC376, rupintrivir, lufotrelvir, PF-07321332, and AG7404.<ref name="Tian_2021">{{cite journal |vauthors =Tian D, Liu Y, Liang C, Xin L, Xie X, Zhang D, Wan M, Li H, Fu X, Liu H, Cao W |title =An update review of emerging small-molecule therapeutic options for COVID-19 |journal =Biomedicine & Pharmacotherapy |volume =137 |date =May 2021 |pmid =33556871 |doi =10.1016/j.biopha.2021.111313 |pmc =7857046 |article-number =111313 }}</ref><ref>{{cite journal |vauthors =Morse JS, Lalonde T, Xu S, Liu WR |title =Learning from the Past: Possible Urgent Prevention and Treatment Options for Severe Acute Respiratory Infections Caused by 2019-nCoV |journal =ChemBioChem |volume =21 |issue =5 |pages =730–738 |date =March 2020 |pmid =32022370 |doi =10.1002/cbic.202000047 |pmc =7162020 |bibcode =2020ChBCh..21..730M }}</ref><ref>{{cite journal |vauthors =Liu C, Zhou Q, Li Y, Garner LV, Watkins SP, Carter LJ, Smoot J, Gregg AC, Daniels AD, Jervey S, Albaiu D |title =Research and Development on Therapeutic Agents and Vaccines for COVID-19 and Related Human Coronavirus Diseases |journal =ACS Central Science |volume =6 |issue =3 |pages =315–331 |date =March 2020 |pmid =32226821 |doi =10.1021/acscentsci.0c00272 |pmc =7094090 |doi-access =free |display-authors =6 |bibcode =2020ACSCS...6..315L }}</ref><ref>{{cite journal |vauthors =Ramajayam R, Tan KP, Liang PH |title =Recent development of 3C and 3CL protease inhibitors for anti-coronavirus and anti-picornavirus drug discovery |journal =Biochemical Society Transactions |volume =39 |issue =5 |pages =1371–5 |date =October 2011 |pmid =21936817 |doi =10.1042/BST0391371 }}</ref><ref name="pmid32321856">{{cite journal |title=Structure-based design of antiviral drug candidates targeting the SARS-CoV-2 main protease |bibcode=2020Sci...368.1331D |doi-access=free |vauthors =Dai W, Zhang B, Jiang XM, Su H, Li J, Zhao Y, Xie X, Jin Z, Peng J, Liu F, Li C, Li Y, Bai F, Wang H, Cheng X, Cen X, Hu S, Yang X, Wang J, Liu X, Xiao G, Jiang H, Rao Z, Zhang LK, Xu Y, Yang H, Liu H |journal =Science |volume =368 |issue =6497 |pages =1331–1335 |date =June 2020 |pmid =32321856 |doi =10.1126/science.abb4489 |pmc =7179937 |display-authors =6}}</ref> The intravenous administered prodrug PF-07304814 (lufotrelvir) entered clinical trials in September 2020.<ref>{{cite journal |author= |date=24 June 2021 |title=First-In-Human Study To Evaluate Safety, Tolerability, And Pharmacokinetics Following Single Ascending And Multiple Ascending Doses of PF-07304814 In Hospitalized Participants With COVID-19. |url=https://clinicaltrials.gov/ct2/show/NCT04535167 |access-date=3 July 2021 |website=Clinical Trials}}</ref>

After clinical trials, in December 2021, the oral medication nirmatrelvir (formerly PF-07321332) became commercially available under emergency use authorizations (EUA), as part of the nirmatrelvir/ritonavir combination therapy (brand name Paxlovid).<ref name="fda-factsheet">{{cite tech report |url=https://www.fda.gov/media/155050/download |title=Fact sheet for healthcare providers: Emergency Use Authorization for Paxlovid |date=22 December 2021 |archive-url=https://web.archive.org/web/20211223210430/https://www.fda.gov/media/155050/download |url-status=live |archive-date=23 December 2021 |format=PDF |publisher=Pfizer |number=LAB-1492-0.8}}</ref><ref name="Pfizer PR 20211222">{{cite press release |title=Pfizer Receives U.S. FDA Emergency Use Authorization for Novel COVID-19 Oral Antiviral Treatment |date=22 December 2021 |publisher=Pfizer |url=https://www.businesswire.com/news/home/20211221005795/en/Pfizer-Receives-U.S.-FDA-Emergency-Use-Authorization-for-Novel-COVID-19-Oral-Antiviral-Treatment |via=Business Wire |access-date=22 December 2021 |url-status=live |archive-url=https://web.archive.org/web/20211222175027/https://www.businesswire.com/news/home/20211221005795/en/Pfizer-Receives-U.S.-FDA-Emergency-Use-Authorization-for-Novel-COVID-19-Oral-Antiviral-Treatment |archive-date=22 December 2021}}</ref> In May 2023, the medication got full FDA approval for high-risk adults, while children 12–18 were still covered under the EUA.<ref name="FDA PR 20230525">{{cite press release |title=FDA Approves First Oral Antiviral for Treatment of COVID-19 in Adults |date=26 May 2023 |url=https://www.fda.gov/news-events/press-announcements/fda-approves-first-oral-antiviral-treatment-covid-19-adults |access-date=26 May 2023 |url-status=dead |archive-url=https://web.archive.org/web/20230525142947/https://www.fda.gov/news-events/press-announcements/fda-approves-first-oral-antiviral-treatment-covid-19-adults |archive-date=May 25, 2023 |website=U.S. Food and Drug Administration (FDA)}} {{PD-notice}}</ref>

The 3C-like protease inhibitor ensitrelvir received authorization to treat COVID-19 in Japan in 2022.<ref name="Shionogi PR 20221122">{{cite press release |title=Xocova (Ensitrelvir Fumaric Acid) Tablets 125mg Approved in Japan for the Treatment of SARS-CoV-2 Infection, under the Emergency Regulatory Approval System |date=22 November 2022 |url=https://www.shionogi.com/us/en/news/2022/11/xocova-ensitrelvir-fumaric-acid-tablets-125mg-approved-in-japan-for-the-treatment-of-sars-cov-2-infection,-under-the-emergency-regulatory-approval-system.html |access-date=28 November 2022 |website=Shionogi}}</ref><ref name="Lenharo 2023">{{cite journal |last=Lenharo |first=Mariana |date=18 October 2023 |title=New Pill Helps COVID Smell and Taste Loss Fade Quickly |url=https://www.scientificamerican.com/article/new-pill-helps-covid-smell-and-taste-loss-fade-quickly/ |journal=Scientific American |doi=10.1038/d41586-023-03244-7 |pmid=37853192 |url-access=subscription |access-date=28 October 2023}}</ref>

In 2022, an ultralarge virtual screening campaign of 235 million molecules was able to identify a novel broad-spectrum inhibitor targeting the main protease of several coronaviruses. It is unusually not a peptidomimetic.<ref name="pmid35142215">{{cite journal |display-authors=6 |bibcode=2022JAChS.144.2905L |vauthors =Luttens A, Gullberg H, Abdurakhmanov E, Vo DD, Akaberi D, Talibov VO, Nekhotiaeva N, Vangeel L, De Jonghe S, Jochmans D, Krambrich J, Tas A, Lundgren B, Gravenfors Y, Craig AJ, Atilaw Y, Sandström A, Moodie LK, Lundkvist Å, van Hemert MJ, Neyts J, Lennerstrand J, Kihlberg J, Sandberg K, Danielson UH, Carlsson J |title =Ultralarge Virtual Screening Identifies SARS-CoV-2 Main Protease Inhibitors with Broad-Spectrum Activity against Coronaviruses |journal =J Am Chem Soc |volume =144 |issue =7 |pages =2905–2920 |date =February 2022 |pmid =35142215 |doi =10.1021/jacs.1c08402 |pmc =8848513 |issn =0002-7863}}</ref>

[[File:Doi.10.1126.science.abb4489.F3.large.C.jpg|thumb|right|A ligand-binding diagram showing the amino acid residues in contact with a covalently bound peptidomimetic protease inhibitor. The small red spheres are water molecules.<ref name="pmid32321856" />]]

== Other 3C(-like) proteases == 3C-like proteases (3C(L)pro) are widely found in (+)ssRNA viruses. All of them are cysteine proteases with a chymotrypsin-like fold (PA clan), using a catalytic dyad or triad. They share some general similarities on substrate specificity and inhibitor effectiveness. They are divided into subfamilies by sequence similarity, corresponding to the family of viruses they are found in:<ref>{{cite journal |vauthors =Kim Y, Lovell S, Tiew KC, Mandadapu SR, Alliston KR, Battaile KP, Groutas WC, Chang KO |title =Broad-spectrum antivirals against 3C or 3C-like proteases of picornaviruses, noroviruses, and coronaviruses |journal =Journal of Virology |volume =86 |issue =21 |pages =11754–62 |date =November 2012 |pmid =22915796 |doi =10.1128/JVI.01348-12 |pmc =3486288 |display-authors =6 }}</ref> * This entry is the coronavirus 3CLpro. * ''Picornaviridae'' have a picornavirus 3Cpro ({{EC number|3.4.22.28}}; {{InterPro|IPR000199}}; MEROPS C03). This is the earliest-studied family. Examples include the ones found in poliovirus and in rhinovirus (both are members of genus ''Enterovirus''). * ''Caliciviridae'' have a 3CLpro ({{InterPro|IPR001665}}; MEROPS C37). Examples include the one found in Norwalk virus.

Additional members are known from ''Potyviridae'' and non-''Coronaviridae'' ''Nidovirales''.<ref>{{cite journal |vauthors =Ziebuhr J, Bayer S, Cowley JA, Gorbalenya AE |title =The 3C-like proteinase of an invertebrate nidovirus links coronavirus and potyvirus homologs |journal =Journal of Virology |volume =77 |issue =2 |pages =1415–26 |date =January 2003 |pmid =12502857 |doi =10.1128/jvi.77.2.1415-1426.2003 |pmc =140795 |doi-access =free }}</ref>

== References == {{reflist}}

== Further reading == {{refbegin}} * {{cite journal |vauthors =Chuck CP, Chow HF, Wan DC, Wong KB |title =Profiling of substrate specificities of 3C-like proteases from group 1, 2a, 2b, and 3 coronaviruses |journal =PLOS ONE |volume =6 |issue =11 |article-number =e27228 |date =2 November 2011 |pmid =22073294 |doi =10.1371/journal.pone.0027228 |pmc =3206940 |doi-access =free |bibcode =2011PLoSO...627228C }} {{refend}}

== External links == * {{MeshName|SARS+coronavirus+main+proteinase}} * Peptidase C30/C16 in coronavirus, {{InterPro|IPR013016}}. The MEROPS C16 one is the "papain-like" PL-PRO.

Category:EC 3.4.22 Category:Coronavirus proteins Category:Viral nonstructural proteins