POLN_EEVV3
ID POLN_EEVV3 Reviewed; 2485 AA.
AC P36327;
DT 01-JUN-1994, integrated into UniProtKB/Swiss-Prot.
DT 10-APR-2019, sequence version 3.
DT 03-AUG-2022, entry version 146.
DE RecName: Full=Polyprotein P1234;
DE Short=P1234;
DE AltName: Full=Non-structural polyprotein;
DE Contains:
DE RecName: Full=Polyprotein P123';
DE Short=P123';
DE Contains:
DE RecName: Full=Polyprotein P123;
DE Short=P123;
DE Contains:
DE RecName: Full=mRNA-capping enzyme nsP1;
DE EC=2.1.1.- {ECO:0000250|UniProtKB:P36328};
DE EC=2.7.7.- {ECO:0000250|UniProtKB:P36328};
DE AltName: Full=Non-structural protein 1;
DE Contains:
DE RecName: Full=Protease nsP2;
DE EC=3.1.3.33 {ECO:0000250|UniProtKB:P08411};
DE EC=3.4.22.- {ECO:0000250|UniProtKB:P27282};
DE EC=3.6.1.15 {ECO:0000250|UniProtKB:Q8JUX6};
DE EC=3.6.4.13 {ECO:0000250|UniProtKB:Q8JUX6};
DE AltName: Full=Non-structural protein 2;
DE Short=nsP2;
DE Contains:
DE RecName: Full=Non-structural protein 3';
DE Short=nsP3';
DE EC=3.1.3.84 {ECO:0000250|UniProtKB:P27282};
DE Contains:
DE RecName: Full=Non-structural protein 3;
DE Short=nsP3;
DE EC=3.1.3.84 {ECO:0000250|UniProtKB:P27282};
DE Contains:
DE RecName: Full=RNA-directed RNA polymerase nsP4;
DE EC=2.7.7.19 {ECO:0000250|UniProtKB:P03317};
DE EC=2.7.7.48 {ECO:0000255|PROSITE-ProRule:PRU00539};
DE AltName: Full=Non-structural protein 4;
DE Short=nsP4;
OS Venezuelan equine encephalitis virus (strain 3880) (VEEV).
OC Viruses; Riboviria; Orthornavirae; Kitrinoviricota; Alsuviricetes;
OC Martellivirales; Togaviridae; Alphavirus.
OX NCBI_TaxID=36382;
OH NCBI_TaxID=9913; Bos taurus (Bovine).
OH NCBI_TaxID=9268; Didelphis marsupialis (Southern opossum).
OH NCBI_TaxID=9793; Equus asinus (Donkey) (Equus africanus asinus).
OH NCBI_TaxID=9796; Equus caballus (Horse).
OH NCBI_TaxID=9606; Homo sapiens (Human).
OH NCBI_TaxID=53535; Melanoconion.
OH NCBI_TaxID=9272; Philander opossum (Gray four-eyed opossum).
OH NCBI_TaxID=10162; Proechimys.
OH NCBI_TaxID=42415; Sigmodon hispidus (Hispid cotton rat).
RN [1]
RP NUCLEOTIDE SEQUENCE [GENOMIC RNA].
RX PubMed=1448915; DOI=10.1016/0042-6822(92)90232-e;
RA Kinney R.M., Tsuchiya K.R., Sneider J.M., Trent D.W.;
RT "Genetic evidence that epizootic Venezuelan equine encephalitis (VEE)
RT viruses may have evolved from enzootic VEE subtype I-D virus.";
RL Virology 191:569-580(1992).
CC -!- FUNCTION: [Polyprotein P1234]: Inactive precursor of the viral
CC replicase, which is activated by cleavages carried out by the viral
CC protease nsP2. {ECO:0000250|UniProtKB:Q8JUX6}.
CC -!- FUNCTION: [Polyprotein P123]: The early replication complex formed by
CC the polyprotein P123 and nsP4 synthesizes the minus-strand RNAs
CC (antigenome) (By similarity). Polyprotein P123 is a short-lived
CC polyprotein that accumulates during early stage of infection
CC (Probable). As soon P123 is cleaved into mature proteins, the plus-
CC strand RNAs synthesis begins (By similarity).
CC {ECO:0000250|UniProtKB:P03317, ECO:0000305}.
CC -!- FUNCTION: [Polyprotein P123']: The early replication complex formed by
CC the polyprotein P123' and nsP4 synthesizes minus-strand RNAs
CC (antigenome) (Probable). Polyprotein P123' is a short-lived polyprotein
CC that accumulates during early stage of infection (Probable). As soon
CC P123' is cleaved into mature proteins, the plus-strand RNAs synthesis
CC begins (Probable). {ECO:0000305}.
CC -!- FUNCTION: [mRNA-capping enzyme nsP1]: Cytoplasmic capping enzyme that
CC catalyzes two virus-specific reactions: methyltransferase and nsP1
CC guanylyltransferase (By similarity). mRNA-capping is necessary since
CC all viral RNAs are synthesized in the cytoplasm, and host capping
CC enzymes are restricted to the nucleus (Probable). The enzymatic
CC reaction involves a covalent link between 7-methyl-GMP and nsP1,
CC whereas eukaryotic capping enzymes form a covalent complex only with
CC GMP (Probable). NsP1 capping consists in the following reactions: GTP
CC is first methylated into 7-methyl-GMP and then is covalently linked to
CC nsP1 to form the m7GMp-nsP1 complex from which 7-methyl-GMP complex is
CC transferred to the mRNA to create the cap structure (By similarity).
CC NsP1 is also needed for the initiation of the minus-strand RNAs
CC synthesis (By similarity). Probably serves as a membrane anchor for the
CC replication complex composed of nsP1-nsP4 (By similarity). Nsp1 is
CC needed for the initiation of the minus-strand RNAs synthesis (By
CC similarity). Palmitoylated nsP1 is remodeling host cell cytoskeleton,
CC and induces filopodium-like structure formation at the surface of the
CC host cell (By similarity). {ECO:0000250|UniProtKB:P03317,
CC ECO:0000250|UniProtKB:P08411, ECO:0000250|UniProtKB:P27282,
CC ECO:0000250|UniProtKB:Q8JUX6, ECO:0000305}.
CC -!- FUNCTION: [Protease nsP2]: Multifunctional protein whose N-terminus is
CC part of the RNA polymerase complex and displays NTPase, RNA
CC triphosphatase and helicase activities (By similarity). NTPase and RNA
CC triphosphatase are involved in viral RNA capping and helicase keeps a
CC check on the dsRNA replication intermediates (By similarity). The C-
CC terminus harbors a protease that specifically cleaves the polyproteins
CC and releases the mature proteins (By similarity). Required for the
CC shutoff of minus-strand RNAs synthesis (By similarity). Inhibits host
CC translation to ensure maximal viral gene expression and evade host
CC immune response (By similarity). {ECO:0000250|UniProtKB:P03317,
CC ECO:0000250|UniProtKB:P08411, ECO:0000250|UniProtKB:P27282,
CC ECO:0000250|UniProtKB:Q8JUX6}.
CC -!- FUNCTION: [Non-structural protein 3]: Seems to be essential for minus-
CC strand RNAs and subgenomic 26S mRNAs synthesis (By similarity).
CC Displays mono-ADP-ribosylhydrolase activity (By similarity). ADP-
CC ribosylation is a post-translational modification that controls various
CC processes of the host cell and the virus probably needs to revert it
CC for optimal viral replication (By similarity). Binds proteins of FXR
CC family and sequesters them into the viral RNA replication complexes
CC thereby inhibiting the formation of host stress granules on viral mRNAs
CC (By similarity). The nsp3-FXR complexes bind viral RNAs and probably
CC orchestrate the assembly of viral replication complexes, thanks to the
CC ability of FXR family members to self-assemble and bind DNA (By
CC similarity). {ECO:0000250|UniProtKB:P03317,
CC ECO:0000250|UniProtKB:P27282}.
CC -!- FUNCTION: [Non-structural protein 3']: Seems to be essential for minus-
CC strand RNAs and subgenomic 26S mRNAs synthesis (By similarity).
CC Displays mono-ADP-ribosylhydrolase activity (Probable). ADP-
CC ribosylation is a post-translational modification that controls various
CC processes of the host cell and the virus probably needs to revert it
CC for optimal viral replication (Probable). Binds proteins of FXR family
CC and sequesters them into the viral RNA replication complexes thereby
CC inhibiting the formation of host stress granules on viral mRNAs
CC (Probable). The nsp3'-FXR complexes bind viral RNAs and probably
CC orchestrate the assembly of viral replication complexes, thanks to the
CC ability of FXR family members to self-assemble and bind DNA (Probable).
CC {ECO:0000250|UniProtKB:P03317, ECO:0000305}.
CC -!- FUNCTION: [RNA-directed RNA polymerase nsP4]: RNA dependent RNA
CC polymerase (By similarity). Replicates genomic and antigenomic RNA by
CC recognizing replications specific signals. The early replication
CC complex formed by the polyprotein P123 and nsP4 synthesizes minus-
CC strand RNAs (By similarity). The late replication complex composed of
CC fully processed nsP1-nsP4 is responsible for the production of genomic
CC and subgenomic plus-strand RNAs (By similarity).
CC {ECO:0000250|UniProtKB:P03317}.
CC -!- CATALYTIC ACTIVITY:
CC Reaction=GTP + S-adenosyl-L-methionine = N(7)-methyl-GTP + S-adenosyl-
CC L-homocysteine; Xref=Rhea:RHEA:46948, ChEBI:CHEBI:37565,
CC ChEBI:CHEBI:57856, ChEBI:CHEBI:59789, ChEBI:CHEBI:87133;
CC Evidence={ECO:0000250|UniProtKB:P27282};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=[nsP1 protein]-L-histidine + N(7)-methyl-GTP = [nsP1 protein]-
CC N(tele)-(N(7)-methylguanosine 5'-phospho)-L-histidine + diphosphate;
CC Xref=Rhea:RHEA:54792, Rhea:RHEA-COMP:13994, Rhea:RHEA-COMP:13995,
CC ChEBI:CHEBI:29979, ChEBI:CHEBI:33019, ChEBI:CHEBI:87133,
CC ChEBI:CHEBI:138334; Evidence={ECO:0000250|UniProtKB:P27282};
CC PhysiologicalDirection=left-to-right; Xref=Rhea:RHEA:54793;
CC Evidence={ECO:0000250|UniProtKB:P27282};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=[nsP1 protein]-N(tele)-(N(7)-methylguanosine 5'-phospho)-L-
CC histidine + a 5'-end diphospho-(purine-ribonucleoside) in mRNA + H(+)
CC = [nsP1 protein]-L-histidine + a 5'-end (N(7)-methyl 5'-
CC triphosphoguanosine)-(purine-ribonucleoside) in mRNA;
CC Xref=Rhea:RHEA:54800, Rhea:RHEA-COMP:12925, Rhea:RHEA-COMP:13929,
CC Rhea:RHEA-COMP:13994, Rhea:RHEA-COMP:13995, ChEBI:CHEBI:15378,
CC ChEBI:CHEBI:29979, ChEBI:CHEBI:133968, ChEBI:CHEBI:138276,
CC ChEBI:CHEBI:138334; Evidence={ECO:0000250|UniProtKB:P27282};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=a 5'-end triphospho-(purine-ribonucleoside) in mRNA + H2O = a
CC 5'-end diphospho-(purine-ribonucleoside) in mRNA + H(+) + phosphate;
CC Xref=Rhea:RHEA:11008, Rhea:RHEA-COMP:13929, Rhea:RHEA-COMP:13942,
CC ChEBI:CHEBI:15377, ChEBI:CHEBI:15378, ChEBI:CHEBI:43474,
CC ChEBI:CHEBI:138276, ChEBI:CHEBI:138288; EC=3.1.3.33;
CC Evidence={ECO:0000250|UniProtKB:P08411};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=a ribonucleoside 5'-triphosphate + H2O = a ribonucleoside 5'-
CC diphosphate + H(+) + phosphate; Xref=Rhea:RHEA:23680,
CC ChEBI:CHEBI:15377, ChEBI:CHEBI:15378, ChEBI:CHEBI:43474,
CC ChEBI:CHEBI:57930, ChEBI:CHEBI:61557; EC=3.6.1.15;
CC Evidence={ECO:0000250|UniProtKB:Q8JUX6};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=ATP + H2O = ADP + H(+) + phosphate; Xref=Rhea:RHEA:13065,
CC ChEBI:CHEBI:15377, ChEBI:CHEBI:15378, ChEBI:CHEBI:30616,
CC ChEBI:CHEBI:43474, ChEBI:CHEBI:456216; EC=3.6.4.13;
CC Evidence={ECO:0000250|UniProtKB:Q8JUX6};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=a ribonucleoside 5'-triphosphate + RNA(n) = diphosphate +
CC RNA(n+1); Xref=Rhea:RHEA:21248, Rhea:RHEA-COMP:14527, Rhea:RHEA-
CC COMP:17342, ChEBI:CHEBI:33019, ChEBI:CHEBI:61557, ChEBI:CHEBI:140395;
CC EC=2.7.7.48; Evidence={ECO:0000255|PROSITE-ProRule:PRU00539};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=4-O-(ADP-D-ribosyl)-L-aspartyl-[protein] + H2O = ADP-D-ribose
CC + H(+) + L-aspartyl-[protein]; Xref=Rhea:RHEA:54428, Rhea:RHEA-
CC COMP:9867, Rhea:RHEA-COMP:13832, ChEBI:CHEBI:15377,
CC ChEBI:CHEBI:15378, ChEBI:CHEBI:29961, ChEBI:CHEBI:57967,
CC ChEBI:CHEBI:138102; Evidence={ECO:0000250|UniProtKB:P27282};
CC PhysiologicalDirection=left-to-right; Xref=Rhea:RHEA:54429;
CC Evidence={ECO:0000250|UniProtKB:P27282};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=5-O-(ADP-D-ribosyl)-L-glutamyl-[protein] + H2O = ADP-D-ribose
CC + H(+) + L-glutamyl-[protein]; Xref=Rhea:RHEA:58248, Rhea:RHEA-
CC COMP:10208, Rhea:RHEA-COMP:15089, ChEBI:CHEBI:15377,
CC ChEBI:CHEBI:15378, ChEBI:CHEBI:29973, ChEBI:CHEBI:57967,
CC ChEBI:CHEBI:142540; Evidence={ECO:0000250|UniProtKB:P27282};
CC PhysiologicalDirection=left-to-right; Xref=Rhea:RHEA:58249;
CC Evidence={ECO:0000250|UniProtKB:P27282};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=ATP + RNA(n) = diphosphate + RNA(n)-3'-adenine ribonucleotide;
CC Xref=Rhea:RHEA:11332, Rhea:RHEA-COMP:14527, Rhea:RHEA-COMP:17347,
CC ChEBI:CHEBI:30616, ChEBI:CHEBI:33019, ChEBI:CHEBI:140395,
CC ChEBI:CHEBI:173115; EC=2.7.7.19;
CC Evidence={ECO:0000250|UniProtKB:P03317};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=ADP-beta-D-ribose 1''-phosphate + H2O = ADP-D-ribose +
CC phosphate; Xref=Rhea:RHEA:25029, ChEBI:CHEBI:15377,
CC ChEBI:CHEBI:43474, ChEBI:CHEBI:57967, ChEBI:CHEBI:58753; EC=3.1.3.84;
CC Evidence={ECO:0000250|UniProtKB:P36328};
CC PhysiologicalDirection=left-to-right; Xref=Rhea:RHEA:25030;
CC Evidence={ECO:0000250|UniProtKB:P36328};
CC -!- COFACTOR:
CC Name=Mg(2+); Xref=ChEBI:CHEBI:18420;
CC Evidence={ECO:0000250|UniProtKB:P03317};
CC Name=Mn(2+); Xref=ChEBI:CHEBI:29035;
CC Evidence={ECO:0000250|UniProtKB:P03317};
CC Note=For nsP4 adenylyltransferase activity; Mn(2+) supports catalysis
CC at 60% of the levels observed with Mg(2+).
CC {ECO:0000250|UniProtKB:P03317};
CC -!- COFACTOR:
CC Name=Mg(2+); Xref=ChEBI:CHEBI:18420;
CC Evidence={ECO:0000250|UniProtKB:P03317};
CC Note=For nsP4 RNA-directed RNA polymerase activity.
CC {ECO:0000250|UniProtKB:P03317};
CC -!- COFACTOR:
CC Name=Mg(2+); Xref=ChEBI:CHEBI:18420;
CC Evidence={ECO:0000250|UniProtKB:P27282};
CC Note=For nsP1 guanylylation. {ECO:0000250|UniProtKB:P27282};
CC -!- COFACTOR:
CC Name=Mg(2+); Xref=ChEBI:CHEBI:18420;
CC Note=For nsP2 RNA triphosphatase activity.
CC {ECO:0000250|UniProtKB:Q8JUX6};
CC -!- COFACTOR:
CC Name=Mg(2+); Xref=ChEBI:CHEBI:18420;
CC Note=For nsP2 NTPase activity. {ECO:0000250|UniProtKB:Q8JUX6};
CC -!- ACTIVITY REGULATION: [mRNA-capping enzyme nsP1]: Inhibited by
CC sinefungin. {ECO:0000250|UniProtKB:P27282}.
CC -!- BIOPHYSICOCHEMICAL PROPERTIES:
CC Kinetic parameters:
CC KM=110 uM for a 12 residues substrate for nsp2 protease activity
CC {ECO:0000250|UniProtKB:P27282};
CC pH dependence:
CC Optimum pH is 7 for nsP1 guanylylation.
CC {ECO:0000250|UniProtKB:P27282};
CC -!- SUBUNIT: [mRNA-capping enzyme nsP1]: Interacts with non-structural
CC protein 3 (By similarity). Interacts with RNA-directed RNA polymerase
CC nsP4 (By similarity). Interacts with protease nsP2 (By similarity).
CC interacts with itself (By similarity). {ECO:0000250|UniProtKB:P27282,
CC ECO:0000250|UniProtKB:Q8JUX6}.
CC -!- SUBUNIT: [Non-structural protein 3]: Interacts with mRNA-capping enzyme
CC nsP1 (By similarity). Interacts with host DDX1 (By similarity).
CC Interacts with host DDX3 (By similarity). Interacts (via C-terminus)
CC with host FXR1; this interaction inhibits the formation of host stress
CC granules on viral mRNAs and the nsp3-FXR1 complexes bind viral RNAs and
CC probably orchestrate the assembly of viral replication complexes (By
CC similarity). Interacts (via C-terminus) with host FXR2; this
CC interaction inhibits the formation of host stress granules on viral
CC mRNAs and the nsp3-FXR2 complexes bind viral RNAs and probably
CC orchestrate the assembly of viral replication complexes (By
CC similarity). Interacts (via C-terminus) with host FMR1; this
CC interaction inhibits the formation of host stress granules on viral
CC mRNAs and the nsp3-FMR1 complexes bind viral RNAs and probably
CC orchestrate the assembly of viral replication complexes (By
CC similarity). {ECO:0000250|UniProtKB:P27282,
CC ECO:0000250|UniProtKB:Q8JUX6}.
CC -!- SUBUNIT: [RNA-directed RNA polymerase nsP4]: Interacts with mRNA-
CC capping enzyme nsP1 (By similarity). Interacts with protease nsP2 (By
CC similarity). interacts with itself (By similarity).
CC {ECO:0000250|UniProtKB:P27282, ECO:0000250|UniProtKB:Q8JUX6}.
CC -!- SUBUNIT: [Protease nsP2]: Interacts with RNA-directed RNA polymerase
CC nsP4 (By similarity). Interacts with mRNA-capping enzyme nsP1 (By
CC similarity). Interacts with KPNA1/karyopherin-alpha1; this interaction
CC probably allows the active transport of protease nsP2 into the host
CC nucleus (By similarity). {ECO:0000250|UniProtKB:P27282,
CC ECO:0000250|UniProtKB:Q8JUX6}.
CC -!- SUBCELLULAR LOCATION: [Polyprotein P1234]: Host cytoplasmic vesicle
CC membrane {ECO:0000305}; Peripheral membrane protein {ECO:0000305}.
CC Note=Part of cytoplasmic vesicles, which are probably formed at the
CC plasma membrane and internalized leading to late endosomal/lysosomal
CC spherules containing the replication complex. {ECO:0000305}.
CC -!- SUBCELLULAR LOCATION: [Polyprotein P123']: Host cytoplasmic vesicle
CC membrane {ECO:0000305}; Peripheral membrane protein {ECO:0000305}.
CC Note=Part of cytoplasmic vesicles, which are probably formed at the
CC plasma membrane and internalized leading to late endosomal/lysosomal
CC spherules containing the replication complex. {ECO:0000305}.
CC -!- SUBCELLULAR LOCATION: [Polyprotein P123]: Host cytoplasmic vesicle
CC membrane {ECO:0000305}; Peripheral membrane protein {ECO:0000305}.
CC Note=Part of cytoplasmic vesicles, which are probably formed at the
CC plasma membrane and internalized leading to late endosomal/lysosomal
CC spherules containing the replication complex. {ECO:0000305}.
CC -!- SUBCELLULAR LOCATION: [mRNA-capping enzyme nsP1]: Host cytoplasmic
CC vesicle membrane {ECO:0000250|UniProtKB:P08411}; Lipid-anchor
CC {ECO:0000250|UniProtKB:P08411}. Host cell membrane
CC {ECO:0000250|UniProtKB:P08411}; Lipid-anchor
CC {ECO:0000250|UniProtKB:P08411}; Cytoplasmic side
CC {ECO:0000250|UniProtKB:P08411}. Host cell projection, host filopodium
CC {ECO:0000250|UniProtKB:P08411}. Note=In the late phase of infection,
CC the polyprotein is quickly cleaved before localization to cellular
CC membranes. Then a fraction of nsP1 localizes to the inner surface of
CC the plasma membrane and its filopodial extensions. Only the
CC palmitoylated nsP1 localizes to the host filopodia (By similarity).
CC NsP1 is also part of cytoplasmic vesicles, which are probably formed at
CC the plasma membrane and internalized leading to late
CC endosomal/lysosomal spherules containing the replication complex (By
CC similarity). {ECO:0000250|UniProtKB:P08411}.
CC -!- SUBCELLULAR LOCATION: [Protease nsP2]: Host cytoplasmic vesicle
CC membrane {ECO:0000250|UniProtKB:P08411}; Peripheral membrane protein
CC {ECO:0000250|UniProtKB:P08411}. Host nucleus
CC {ECO:0000250|UniProtKB:P27282}. Host cytoplasm
CC {ECO:0000250|UniProtKB:P27282}. Note=In the late phase of infection,
CC the polyprotein is quickly cleaved before localization to cellular
CC membranes. Then approximately half of nsP2 is found in the nucleus (By
CC similarity). Shuttles between cytoplasm and nucleus (By similarity).
CC NsP2 is also part of cytoplasmic vesicles, which are probably formed at
CC the plasma membrane and internalized leading to late
CC endosomal/lysosomal spherules containing the replication complex (By
CC similarity). {ECO:0000250|UniProtKB:P08411,
CC ECO:0000250|UniProtKB:P27282}.
CC -!- SUBCELLULAR LOCATION: [Non-structural protein 3]: Host cytoplasmic
CC vesicle membrane {ECO:0000250|UniProtKB:P03317}; Peripheral membrane
CC protein {ECO:0000305}. Note=In the late phase of infection, the
CC polyprotein is quickly cleaved before localization to cellular
CC membranes. Then nsP3 and nsP3' form aggregates in cytoplasm (By
CC similarity). NsP3 is also part of cytoplasmic vesicles, which are
CC probably formed at the plasma membrane and internalized leading to late
CC endosomal/lysosomal spherules containing the replication complex (By
CC similarity). {ECO:0000250|UniProtKB:P03317}.
CC -!- SUBCELLULAR LOCATION: [Non-structural protein 3']: Host cytoplasmic
CC vesicle membrane {ECO:0000305}; Peripheral membrane protein
CC {ECO:0000305}. Note=In the late phase of infection, the polyprotein is
CC quickly cleaved before localization to cellular membranes. Then nsP3
CC and nsP3' form aggregates in cytoplasm (By similarity). NsP3' is also
CC part of cytoplasmic vesicles, which are probably formed at the plasma
CC membrane and internalized leading to late endosomal/lysosomal spherules
CC containing the replication complex (Probable).
CC {ECO:0000250|UniProtKB:P03317, ECO:0000305}.
CC -!- SUBCELLULAR LOCATION: [RNA-directed RNA polymerase nsP4]: Host
CC cytoplasmic vesicle membrane; Peripheral membrane protein
CC {ECO:0000250|UniProtKB:P08411}. Note=NsP4 is part of cytoplasmic
CC vesicles, which are probably formed at the plasma membrane and
CC internalized leading to late endosomal/lysosomal spherules containing
CC the replication complex. {ECO:0000250|UniProtKB:P08411}.
CC -!- DOMAIN: [Protease nsP2]: The N-terminus exhibits NTPase and RNA
CC triphosphatase activities and is proposed to have helicase activity,
CC whereas the C-terminus possesses protease activity (By similarity).
CC Contains a nuclear localization signal and a nuclear export signal,
CC these two motifs are probably involved in the shuttling between the
CC cytoplasm and the nucleus of nsP2 (By similarity).
CC {ECO:0000250|UniProtKB:P27282, ECO:0000250|UniProtKB:Q8JUX6}.
CC -!- DOMAIN: [Non-structural protein 3]: In the N-terminus, the macro domain
CC displays a mono-ADP-ribosylhydrolase activity (By similarity). The
CC central part has a zinc-binding function (By similarity). The C-
CC terminus contains two approximate repeats necessary and sufficient for
CC formation of the nsP3/FXR complex (By similarity).
CC {ECO:0000250|UniProtKB:P03317, ECO:0000250|UniProtKB:P27282}.
CC -!- DOMAIN: [Non-structural protein 3']: In the N-terminus, the macro
CC domain displays a mono-ADP-ribosylhydrolase activity (By similarity).
CC The central part has a zinc-binding function (By similarity). The C-
CC terminus contains two approximate repeats necessary and sufficient for
CC formation of the nsP3'/FXR complex (By similarity).
CC {ECO:0000250|UniProtKB:P03317, ECO:0000250|UniProtKB:P27282}.
CC -!- PTM: [Polyprotein P1234]: Specific enzymatic cleavages in vivo yield
CC mature proteins (By similarity). The processing of the polyprotein is
CC temporally regulated (By similarity). In early stages (1.7 hpi), P1234
CC is first cleaved in trans through its nsP2 protease activity, releasing
CC P123' and nsP4, which associate to form the early replication complex
CC (By similarity). At the same time, P1234 is also cut at the nsP1/nsP2
CC site early in infection but with lower efficiency (By similarity).
CC After replication of the viral minus-strand RNAs (4 hpi), the
CC polyproteins are cut at the nsP1/nsP2 and nsP2/nsP3 sites very
CC efficiently, preventing accumulation of P123' and P1234 and allowing
CC the formation of the late replication complex (By similarity).
CC NsP3'/nsP4 site is not cleaved anymore and P34 is produced rather than
CC nsP4 (By similarity). {ECO:0000250|UniProtKB:P03317}.
CC -!- PTM: [Polyprotein P123]: Specific enzymatic cleavages in vivo yield
CC mature proteins (By similarity). The processing of the polyprotein is
CC temporally regulated (By similarity). In early stages (1.7 hpi), P123
CC is cleaved at the nsP1/nsP2 site with low efficiency (By similarity).
CC After replication of the viral minus-strand RNAs (4 hpi), the
CC polyproteins are cut at the nsP1/nsP2 and nsP2/nsP3 sites very
CC efficiently, preventing accumulation of P123 and allowing the formation
CC of the late replication complex (By similarity).
CC {ECO:0000250|UniProtKB:P03317}.
CC -!- PTM: [Polyprotein P123']: Specific enzymatic cleavages in vivo yield
CC mature proteins (By similarity). The processing of the polyprotein is
CC temporally regulated (By similarity). In early stages (1.7 hpi), P123'
CC is cleaved at the nsP1/nsP2 site with low efficiency (By similarity).
CC After replication of the viral minus-strand RNAs (4 hpi), the
CC polyproteins are cut at the nsP1/nsP2 and nsP2/nsP3 sites very
CC efficiently, preventing accumulation of P123' and allowing the
CC formation of the late replication complex (By similarity).
CC {ECO:0000250|UniProtKB:P03317}.
CC -!- PTM: [mRNA-capping enzyme nsP1]: Palmitoylated by host
CC palmitoyltransferases ZDHHC2 and ZDHHC19.
CC {ECO:0000250|UniProtKB:P03317}.
CC -!- PTM: [Non-structural protein 3]: Phosphorylated by host on serines and
CC threonines. {ECO:0000250|UniProtKB:P08411}.
CC -!- PTM: [Non-structural protein 3']: Phosphorylated by host on serines and
CC threonines. {ECO:0000250|UniProtKB:P08411}.
CC -!- PTM: [RNA-directed RNA polymerase nsP4]: Ubiquitinated; targets the
CC protein for rapid degradation via the ubiquitin system (By similarity).
CC Nsp4 is present in extremely low quantities due to low frequency of
CC translation through the amber stop-codon and the degradation by the
CC ubiquitin pathway (By similarity). {ECO:0000250|UniProtKB:P03317}.
CC -!- MISCELLANEOUS: Viral replication produces dsRNA in the late phase of
CC infection, resulting in a strong activation of host EIF2AK2/PKR,
CC leading to almost complete phosphorylation of EIF2A (By similarity).
CC This inactivates completely cellular translation initiation, resulting
CC shutoff of host proteins synthesis (By similarity). However,
CC phosphorylation of EIF2A is probably not the only mechanism responsible
CC for the host translation shutoff (By similarity). The viral translation
CC can still occur normally because it relies on a hairpin structure in
CC the coding region of sgRNA and is EIF2A-, EIF2D-, EIF4G- EIF4A-
CC independent (By similarity). {ECO:0000250|UniProtKB:P03317}.
CC -!- MISCELLANEOUS: The genome codes for P123, but readthrough of a
CC terminator codon UGA occurs between the codons for Gln-1871 and Arg-
CC 1873 giving rise to P1234. P1234 is cleaved quickly by nsP2 into P123'
CC and nsP4 (By similarity). Further processing of p123' gives nsP1, nsP2
CC and nsP3' which is 6 amino acids longer than nsP3 since the cleavage
CC site is after the readthrough (By similarity). This unusual molecular
CC mechanism ensures that few nsP4 are produced compared to other non-
CC structural proteins (By similarity). Mutant viruses with no alternative
CC termination site grow significantly slower than wild-type virus (By
CC similarity). The opal termination codon is frequently mutated to a
CC sense codon on passage in cell culture (By similarity). The presence of
CC the opal codon may be a requirement for viral maintenance in both
CC vertebrate and invertebrate hosts and a selective advantage may be
CC conferred in cell culture for the sense codon (By similarity).
CC {ECO:0000250|UniProtKB:O90368, ECO:0000250|UniProtKB:P03317}.
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DR EMBL; L00930; AAC19324.1; -; Genomic_RNA.
DR EMBL; L00930; AAC19326.1; ALT_SEQ; Genomic_RNA.
DR PIR; C44213; C44213.
DR MEROPS; C09.002; -.
DR PRIDE; P36327; -.
DR Proteomes; UP000008299; Genome.
DR GO; GO:0044162; C:host cell cytoplasmic vesicle membrane; IEA:UniProtKB-SubCell.
DR GO; GO:0044176; C:host cell filopodium; IEA:UniProtKB-SubCell.
DR GO; GO:0042025; C:host cell nucleus; IEA:UniProtKB-SubCell.
DR GO; GO:0020002; C:host cell plasma membrane; IEA:UniProtKB-SubCell.
DR GO; GO:0016020; C:membrane; IEA:UniProtKB-KW.
DR GO; GO:0005524; F:ATP binding; IEA:UniProtKB-KW.
DR GO; GO:0016887; F:ATP hydrolysis activity; IEA:RHEA.
DR GO; GO:0008234; F:cysteine-type peptidase activity; IEA:UniProtKB-KW.
DR GO; GO:0005525; F:GTP binding; IEA:UniProtKB-KW.
DR GO; GO:0046872; F:metal ion binding; IEA:UniProtKB-KW.
DR GO; GO:0008174; F:mRNA methyltransferase activity; IEA:InterPro.
DR GO; GO:0004651; F:polynucleotide 5'-phosphatase activity; IEA:UniProtKB-EC.
DR GO; GO:0004652; F:polynucleotide adenylyltransferase activity; IEA:UniProtKB-EC.
DR GO; GO:0003723; F:RNA binding; IEA:UniProtKB-KW.
DR GO; GO:0003724; F:RNA helicase activity; IEA:UniProtKB-EC.
DR GO; GO:0003968; F:RNA-directed 5'-3' RNA polymerase activity; IEA:UniProtKB-KW.
DR GO; GO:0006370; P:7-methylguanosine mRNA capping; IEA:UniProtKB-KW.
DR GO; GO:0006508; P:proteolysis; IEA:UniProtKB-KW.
DR GO; GO:0039523; P:suppression by virus of host mRNA transcription via inhibition of RNA polymerase II activity; IEA:UniProtKB-KW.
DR GO; GO:0006351; P:transcription, DNA-templated; IEA:InterPro.
DR GO; GO:0039694; P:viral RNA genome replication; IEA:InterPro.
DR CDD; cd21557; Macro_X_Nsp3-like; 1.
DR Gene3D; 3.40.220.10; -; 1.
DR Gene3D; 3.40.50.150; -; 1.
DR Gene3D; 3.40.50.300; -; 2.
DR Gene3D; 3.90.70.110; -; 1.
DR InterPro; IPR027351; (+)RNA_virus_helicase_core_dom.
DR InterPro; IPR002588; Alphavirus-like_MT_dom.
DR InterPro; IPR002620; Alphavirus_nsp2pro.
DR InterPro; IPR044936; Alphavirus_nsp2pro_sf.
DR InterPro; IPR043502; DNA/RNA_pol_sf.
DR InterPro; IPR002589; Macro_dom.
DR InterPro; IPR043472; Macro_dom-like.
DR InterPro; IPR044371; Macro_X_NSP3-like.
DR InterPro; IPR027417; P-loop_NTPase.
DR InterPro; IPR007094; RNA-dir_pol_PSvirus.
DR InterPro; IPR029063; SAM-dependent_MTases_sf.
DR InterPro; IPR001788; Tymovirus_RNA-dep_RNA_pol.
DR Pfam; PF01661; Macro; 1.
DR Pfam; PF01707; Peptidase_C9; 1.
DR Pfam; PF00978; RdRP_2; 1.
DR Pfam; PF01443; Viral_helicase1; 1.
DR Pfam; PF01660; Vmethyltransf; 1.
DR SMART; SM00506; A1pp; 1.
DR SUPFAM; SSF52540; SSF52540; 1.
DR SUPFAM; SSF52949; SSF52949; 1.
DR SUPFAM; SSF56672; SSF56672; 1.
DR PROSITE; PS51743; ALPHAVIRUS_MT; 1.
DR PROSITE; PS51154; MACRO; 1.
DR PROSITE; PS51520; NSP2PRO; 1.
DR PROSITE; PS51657; PSRV_HELICASE; 1.
DR PROSITE; PS50507; RDRP_SSRNA_POS; 1.
PE 3: Inferred from homology;
KW ATP-binding; Eukaryotic host gene expression shutoff by virus;
KW Eukaryotic host transcription shutoff by virus; GTP-binding; Helicase;
KW Host cell membrane; Host cell projection; Host cytoplasm;
KW Host cytoplasmic vesicle; Host gene expression shutoff by virus;
KW Host membrane; Host nucleus; Host-virus interaction; Hydrolase;
KW Inhibition of host RNA polymerase II by virus; Lipoprotein; Membrane;
KW Metal-binding; Methyltransferase; mRNA capping; mRNA processing;
KW Multifunctional enzyme; Nucleotide-binding; Nucleotidyltransferase;
KW Palmitate; Phosphoprotein; Protease; Repeat;
KW RNA suppression of termination; RNA-binding; RNA-directed RNA polymerase;
KW S-adenosyl-L-methionine; Thiol protease; Transferase; Ubl conjugation;
KW Viral RNA replication; Zinc.
FT CHAIN 1..2485
FT /note="Polyprotein P1234"
FT /id="PRO_0000308387"
FT CHAIN 1..1878
FT /note="Polyprotein P123'"
FT /id="PRO_0000228750"
FT CHAIN 1..1871
FT /note="Polyprotein P123"
FT /id="PRO_0000228751"
FT CHAIN 1..535
FT /note="mRNA-capping enzyme nsP1"
FT /id="PRO_0000041200"
FT CHAIN 536..1329
FT /note="Protease nsP2"
FT /id="PRO_0000041201"
FT CHAIN 1330..1878
FT /note="Non-structural protein 3'"
FT /id="PRO_0000228752"
FT CHAIN 1330..1871
FT /note="Non-structural protein 3"
FT /id="PRO_0000041202"
FT CHAIN 1879..2485
FT /note="RNA-directed RNA polymerase nsP4"
FT /id="PRO_0000041203"
FT DOMAIN 28..259
FT /note="Alphavirus-like MT"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU01079"
FT DOMAIN 690..841
FT /note="(+)RNA virus helicase ATP-binding"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU00990"
FT DOMAIN 842..990
FT /note="(+)RNA virus helicase C-terminal"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU00990"
FT DOMAIN 1003..1322
FT /note="Peptidase C9"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU00853"
FT DOMAIN 1328..1489
FT /note="Macro"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU00490"
FT REPEAT 1810..1831
FT /note="1"
FT /evidence="ECO:0000250|UniProtKB:P27282"
FT REPEAT 1844..1865
FT /note="2"
FT /evidence="ECO:0000250|UniProtKB:P27282"
FT DOMAIN 2242..2357
FT /note="RdRp catalytic"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU00539"
FT REGION 244..263
FT /note="NsP1 membrane-binding"
FT /evidence="ECO:0000250|UniProtKB:P08411"
FT REGION 1004..1023
FT /note="Nucleolus localization signal"
FT /evidence="ECO:0000250|UniProtKB:P08411"
FT REGION 1783..1810
FT /note="Disordered"
FT /evidence="ECO:0000256|SAM:MobiDB-lite"
FT REGION 1810..1865
FT /note="2 X 21 AA approximate repeats, binding to host FXR
FT family members"
FT /evidence="ECO:0000250|UniProtKB:P27282"
FT MOTIF 1056..1065
FT /note="Nuclear export signal"
FT /evidence="ECO:0000250|UniProtKB:P27282"
FT MOTIF 1179..1183
FT /note="Nuclear localization signal"
FT /evidence="ECO:0000250|UniProtKB:P27282"
FT COMPBIAS 1784..1798
FT /note="Pro residues"
FT /evidence="ECO:0000256|SAM:MobiDB-lite"
FT ACT_SITE 1012
FT /note="For cysteine protease nsP2 activity"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU00853"
FT ACT_SITE 1081
FT /note="For cysteine protease nsP2 activity"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU00853"
FT BINDING 721..728
FT /ligand="a ribonucleoside 5'-triphosphate"
FT /ligand_id="ChEBI:CHEBI:61557"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU00990"
FT BINDING 1339
FT /ligand="ADP-D-ribose"
FT /ligand_id="ChEBI:CHEBI:57967"
FT /evidence="ECO:0000250|UniProtKB:P36328"
FT BINDING 1353
FT /ligand="ADP-D-ribose"
FT /ligand_id="ChEBI:CHEBI:57967"
FT /evidence="ECO:0000250|UniProtKB:Q8JUX6"
FT BINDING 1361
FT /ligand="ADP-D-ribose"
FT /ligand_id="ChEBI:CHEBI:57967"
FT /evidence="ECO:0000250|UniProtKB:Q8JUX6"
FT BINDING 1441
FT /ligand="ADP-D-ribose"
FT /ligand_id="ChEBI:CHEBI:57967"
FT /evidence="ECO:0000250|UniProtKB:P36328"
FT BINDING 1442
FT /ligand="ADP-D-ribose"
FT /ligand_id="ChEBI:CHEBI:57967"
FT /evidence="ECO:0000250|UniProtKB:P36328"
FT BINDING 1443
FT /ligand="ADP-D-ribose"
FT /ligand_id="ChEBI:CHEBI:57967"
FT /evidence="ECO:0000250|UniProtKB:P36328"
FT BINDING 1596
FT /ligand="Zn(2+)"
FT /ligand_id="ChEBI:CHEBI:29105"
FT /evidence="ECO:0000250|UniProtKB:P03317"
FT BINDING 1598
FT /ligand="Zn(2+)"
FT /ligand_id="ChEBI:CHEBI:29105"
FT /evidence="ECO:0000250|UniProtKB:P03317"
FT BINDING 1621
FT /ligand="Zn(2+)"
FT /ligand_id="ChEBI:CHEBI:29105"
FT /evidence="ECO:0000250|UniProtKB:P03317"
FT BINDING 1639
FT /ligand="Zn(2+)"
FT /ligand_id="ChEBI:CHEBI:29105"
FT /evidence="ECO:0000250|UniProtKB:P03317"
FT SITE 37
FT /note="Involved in the phosphoramide link with 7-methyl-
FT GMP"
FT /evidence="ECO:0000250|UniProtKB:P27282"
FT SITE 535..536
FT /note="Cleavage; by protease nsP2"
FT /evidence="ECO:0000250|UniProtKB:P03317"
FT SITE 1329..1330
FT /note="Cleavage; by protease nsP2"
FT /evidence="ECO:0000250|UniProtKB:P03317"
FT SITE 1878..1879
FT /note="Cleavage; by protease nsP2"
FT /evidence="ECO:0000250|UniProtKB:Q8JUX6"
FT LIPID 419
FT /note="S-palmitoyl cysteine; by host"
FT /evidence="ECO:0000250|UniProtKB:Q8JUX6"
SQ SEQUENCE 2485 AA; 277240 MW; 328AB6665D045459 CRC64;
MEKVHVDIEE DSPFLRALQR SFPQFEVEAK QVTDNDHANA RAFSHLASKL IETEVDPSDT
ILDIGSAPAR RMYSKHKYHC ICPMRCAEDP DRLYKYATKL KKNCKEITDK ELDKKMKELA
AVMSDPDLET ETMCLHDDES CRYEGQVAVY QDVYAVDGPT SLYHQANKGV RVAYWIGFDT
TPFMFKNLAG AYPSYSTNWA DETVLTARNI GLCSSDVMER SRRGMSILRK KYLKPSNNVL
FSVGSTIYHE KRDLLRSWHL PSVFHLRGKQ NYTCRCETIV SCDGYVVKRI AISPGLYGKP
SGYAATMHRE GFLCCKVTDT LNGERVSFPV CTYVPATLCD QMTGILATDV SADDAQKLLV
GLNQRIVVNG RTQRNTNTMK NYLLPVVAQA FARWAKEYKE DQEDERPLGL RDRQLVMGCC
WAFRRHKITS IYKRPDTQTI IKVNSDFHSF VLPRIGSNTL EIGLRTRIRK MLEEHKEPSP
LITAEDIQEA KCAADEAKEV REAEELRAVL PPLAADVEEP TLEADVDLML QEAGAGSVET
PRGLIKVTSY AGEDKIGSYA VLSPQAVLKS EKLSCIHPLA EQVIVITHSG RKGRYAVEPY
HGKVVVPEGH AIPVQDFQAL SESATIVYNE REFVNRYLHH IATHGGALNT DEEYYKTVKP
SEHDGEYLYD IDRKQCVKKE LVTGLGLTGE LVDPPFHEFA YESLRTRPAA PYQVPTIGVY
GVPGSGKSGI IKSAVTKKDL VVSAKKENCA EIIRDVKRIK GLDVNARTVD SVLLNGCKYP
VETLYIDEAF ACHAGTLRAL IAIIRPKKAV LCGDPKQCGF FNMMCLKVHF NHEICTQVFH
KSISRRCTKS VTSVVSTLFY DKRMRTTNPK ETKIEIDTTG STKPKQDDLI LTCFRGWVKQ
LQIDYKGNEV MTAAASQGLT RKGVYAVRYK VNENPLYAPT SEHVNVLLTR TEDRIVWKTL
AGDPWIKTLT AKYPGNFTAT IEEWQAEHDA IMRHILERPD PTDVFQNKAN VCWAKALVPV
LKTAGIDMTT EQWNTVDYFE TDKAHSAEIV LNQLCVRFFG LDLDSGLFSA PTVPLSIRNN
HWDNSPSPNM YGLNKEVVRQ LSRRYPQLPR AVTTGRVYDM NTGTLRNYDP RINLVPVNRR
LPHALVLHHN EHPQSDFSSF VSKLKGRTVL VVGEKLSVPG KTVDWLSDRP EATFRARLDL
GIPGDVPKYD IIFINVRTPY KYHHYQQCED HAIKLSMLTK KACLHLNPGG TCVSIGYGYA
DRASESIIGA IARQFKFSRV CKPKSSLEET EVLFVFIGYD RKARTHNPYK LSSTLTNIYT
GSRLHEAGCA PSYHVVRGDI ATATEGVIIN AANSKGQPGG GVCGALYKKF PESFDLQPIE
VGKARLVKGA AKHIIHAVGP NFNKVSEIEG DKQLAEAYES IAKIVNDNNY KSVAIPLLST
GIFSGNKDRL TQSLNHLLTA LDTTDADVAI YCRDKKWEMT LKEAVARREA VEEICISDDS
SVTEPDAELV RVHPKSSLAG RKGYSTSDGK TFSYLEGTKF HQAAKDIAEI NAMWPVATEA
NEQVCMYILG ESMSSIRSKC PVEESEASTP PSTLPCLCIH AMTPERVQRL KASRPEQITV
CSSFPLPKYR ITGVQKIQCS QPILFSPKVP AYIHPRKYLV ETPTVEENQS TEGTPEQPTL
ITVGETRTRT PEPIIIEEEE DSISLLSDGP THQVLQVEAD IHGPPSASSS SWSIPHASDF
DVDSLSILDT LEGASVTSEE ASVETNSHFA RSMEFLARPV PAPRTVFRNP PQPAPRTRTP
SLAPSRASSR ISLVSNPPGV NRVITREELE ALTPSRAPSR SVSRTSLVSN PPGVNRVITR
EEFEAFVAQQ QXRFDAGAYI FSSDTGQGHL QQKSVRQTVL SEVVLERTEL EISYAPRLDQ
EKEELLRKKL QLNPTPANRS RYQSRRVENM KAITARRILQ GLGHYLKAEG KVECYRTLHP
VPLYSSSVNR AFSSPKVAVE ACNAMLKENF PTVASYCIIP EYDAYLDMVD GASCCLDTAS
FCPAKLRSFP KKHSYLEPTI RSAVPSAIQN TLQNVLAAAT KRNCNVTQMR ELPVLDSAAF
NVECFKKYAC NNEYWETFKE NPIRLTEENV VNYITKLKGP KAAALFAKTH NLNMLQDIPM
DRFVMDLKRD VKVTPGTKHT EERPKVQVIQ AADPLATAYL CGIHRELVRR LNAVLLPNIH
TLFDMSAEDF DAIIAEHFQP GDCVLETDIA SFDKSEDDAM ALTALMILED LGVDAELLTL
IEAAFGEISS IHLPTKTKFK FGAMMKSGMF LTLFVNTVIN IVIASRVLRE RLTGSPCAAF
IGDDNIVKGV KSDKLMADRC ATWLNMEVKI IDAVVGEKAP YFCGGFILCD SVTGTACRVA
DPLKRLFKLG KPLAADDEHD DDRRRALHEE STRWNRVGIL PELCKAVESR YETVGTSIIV
MAMTTLASSV KSFSYLRGAP ITLYG
