IntAct European Bioinformatics Institute IntAct intact http://www.ebi.ac.uk/ http://www.ebi.ac.uk/intact/query/${ac} EBI-[0-9]+|IA:[0-9]+ INTerAction database (IntAct) provides an open source database and toolkit for the storage, presentation and analysis of molecular interactions. http://www.ebi.ac.uk/intact European Bioinformatics Institute; Wellcome Trust Genome Campus; Hinxton, Cambridge; CB10 1SD; United Kingdom http://www.ebi.ac.uk/intact/ Short peptides derived from the interaction domain of SARS coronavirus nonstructural protein nsp10 can suppress the 2'-O-methyltransferase activity of nsp10/nsp16 complex. Short peptides derived from the interaction domain of SARS coronavirus nonstructural protein nsp10 can suppress the 2'-O-methyltransferase activity of nsp10/nsp16 complex. Virus research 2012 imex curation Ke M., Chen Y., Wu A., Sun Y., Su C., Wu H., Jin X., Tao J., Wang Y., Ma X., Pan JA., Guo D. Virus - Publications including interactions involving viral proteins Coronavirus - Interactions investigated in the context of Coronavirus dguo@whu.edu.cn 28-Sep-2020: Contacted by IntAct-Help. Only protein-protein interactions imex curation yeasx Saccharomyces cerevisiae (Baker's yeast) Baker's yeast 2 hybrid two hybrid 2H 2h Gal4 transcription regeneration two-hybrid yeast two hybrid 2-hybrid classical two hybrid classical two hybrid 2H two-hybrid 2-hybrid Gal4 transcription regeneration 2h yeast two hybrid Y2H Y-2H 2 hybrid predetermined predetermined participant predetermined Ke M., Chen Y., Wu A., Sun Y., Su C., Wu H., Jin X., Tao J., Wang Y., Ma X., Pan JA., Guo D. Virus - Publications including interactions involving viral proteins dguo@whu.edu.cn Accepted 2020-AUG-19 AT 10:07 BST AT 10:07 BST by MARTA Virus research 2012 Only protein-protein interactions imex curation imex curation Short peptides derived from the interaction domain of SARS coronavirus nonstructural protein nsp10 can suppress the 2'-O-methyltransferase activity of nsp10/nsp16 complex. Short peptides derived from the interaction domain of SARS coronavirus nonstructural protein nsp10 can suppress the 2'-O-methyltransferase activity of nsp10/nsp16 complex. Virus research 2012 imex curation Ke M., Chen Y., Wu A., Sun Y., Su C., Wu H., Jin X., Tao J., Wang Y., Ma X., Pan JA., Guo D. Virus - Publications including interactions involving viral proteins Coronavirus - Interactions investigated in the context of Coronavirus dguo@whu.edu.cn 28-Sep-2020: Contacted by IntAct-Help. Only protein-protein interactions imex curation in vitro In vitro methyltransferase as methyltransferase assay methyltransferase as tag visualisation tag visualisation Ke M., Chen Y., Wu A., Sun Y., Su C., Wu H., Jin X., Tao J., Wang Y., Ma X., Pan JA., Guo D. Virus - Publications including interactions involving viral proteins Radiolabeled S-adenosyl [methyl-3H] methionine, purified SARS-CoV nsp16/nsp10 complex, +/- short peptides, was mixed with m7GpppA-RNA substrate. The 3H-labeled product was isolated in small DEAE-Sephadex columns and quantitated by liquid scintillation counting. dguo@whu.edu.cn Accepted 2020-AUG-19 AT 10:07 BST AT 10:07 BST by MARTA Virus research 2012 Only protein-protein interactions imex curation imex curation Short peptides derived from the interaction domain of SARS coronavirus nonstructural protein nsp10 can suppress the 2'-O-methyltransferase activity of nsp10/nsp16 complex. Short peptides derived from the interaction domain of SARS coronavirus nonstructural protein nsp10 can suppress the 2'-O-methyltransferase activity of nsp10/nsp16 complex. Virus research 2012 imex curation Ke M., Chen Y., Wu A., Sun Y., Su C., Wu H., Jin X., Tao J., Wang Y., Ma X., Pan JA., Guo D. Virus - Publications including interactions involving viral proteins Coronavirus - Interactions investigated in the context of Coronavirus dguo@whu.edu.cn 28-Sep-2020: Contacted by IntAct-Help. Only protein-protein interactions imex curation in vitro In vitro methyltransferase as methyltransferase assay methyltransferase as autoradiography autoradiography Ke M., Chen Y., Wu A., Sun Y., Su C., Wu H., Jin X., Tao J., Wang Y., Ma X., Pan JA., Guo D. Virus - Publications including interactions involving viral proteins Reaction mixtures were spotted onto TLC plates. Radiolabelling was determined using a PhosphorImager. dguo@whu.edu.cn Accepted 2020-AUG-19 AT 10:07 BST AT 10:07 BST by MARTA Virus research 2012 Only protein-protein interactions imex curation imex curation Short peptides derived from the interaction domain of SARS coronavirus nonstructural protein nsp10 can suppress the 2'-O-methyltransferase activity of nsp10/nsp16 complex. Short peptides derived from the interaction domain of SARS coronavirus nonstructural protein nsp10 can suppress the 2'-O-methyltransferase activity of nsp10/nsp16 complex. Virus research 2012 imex curation Ke M., Chen Y., Wu A., Sun Y., Su C., Wu H., Jin X., Tao J., Wang Y., Ma X., Pan JA., Guo D. Virus - Publications including interactions involving viral proteins Coronavirus - Interactions investigated in the context of Coronavirus dguo@whu.edu.cn 28-Sep-2020: Contacted by IntAct-Help. Only protein-protein interactions imex curation in vitro In vitro crosslink cross-linking study crosslink autoradiography autoradiography Ke M., Chen Y., Wu A., Sun Y., Su C., Wu H., Jin X., Tao J., Wang Y., Ma X., Pan JA., Guo D. Virus - Publications including interactions involving viral proteins SAM binding assay:protein mixtures incubated with S-adenosyl [methyl-3H] methionine were irradiated with UV light to cross-link SAM to its binding site. Samples were separated by SDS-PAGE and and analyzed by autoradiography. dguo@whu.edu.cn Accepted 2020-AUG-19 AT 10:07 BST AT 10:07 BST by MARTA Virus research 2012 Only protein-protein interactions imex curation imex curation sars_rna_substrate rna ribonucleic acid RNA rna synthetic construct synthetic construct AGAUUAGGUUUUUACCUACCCAGGAAAAGCCAACCAACCUCGAUCUCUUGUAGAUCUGUUCUCUAAACGAACUUUAAAAUCUGUGUAGCUGUCGCUCGGCUGCAUGCCUAGUGCACCUACGCAGUAUAAACAAUAAUAAAUUUUACUGUCGUUGACAAGAAACGAGUAACUCGUCCCUCUUCUGCAGACUGCUUACGGUUUCGUCCGUGUUGCAGUCGAUCAUCAGCAUACCUAGGUUUCGUCCGGGUGUGACCGAAAG 5′-terminal 259 nucleotides of the SARS-CoV genome, strain WHU, GenBank: AY394850.2, with first U mutated to a G (pmid:19208801) p0c6x7-pro_0000037320 Proofreading exoribonuclease nsp14 rep ORF1ab polyprotein 1a-1b protein protein sars-cov Human SARS coronavirus SARS-CoV Severe acute respiratory syndrome coronavirus AENVTGLFKDCSKIITGLHPTQAPTHLSVDIKFKTEGLCVDIPGIPKDMTYRRLISMMGFKMNYQVNGYPNMFITREEAIRHVRAWIGFDVEGCHATRDAVGTNLPLQLGFSTGVNLVAVPTGYVDTENNTEFTRVNAKPPPGDQFKHLIPLMYKGLPWNVVRIKIVQMLSDTLKGLSDRVVFVLWAHGFELTSMKYFVKIGPERTCCLCDKRATCFSTSSDTYACWNHSVGFDYVYNPFMIDVQQWGFTGNLQSNHDQHCQVHGNAHVASCDAIMTRCLAVHECFVKRVDWSVEYPIIGDELRVNSACRKVQHMVVKSALLADKFPVLHDIGNPKAIKCVPQAEVEWKFYDAQPCSDKAYKIEELFYSYATHHDKFTDGVCLFWNCNVDRYPANAIVCRFDTRVLSNLNLPGCDGGSLYVNKHAFHTPAFDKSAFTNLKQLPFFYYSDSPCESHGKQVVSDIDYVPLKSATCITRCNLGGAVCRHHANEYRQYLDAYNMMISAGFSLWIYKQFDTYNLWNTFTRLQ 5903 6429 D8E25737EF40967D p0c6x7-pro_0000037322 2'-O-methyltransferase nsp16 ORF1ab polyprotein 1a-1b rep protein protein sars-cov Human SARS coronavirus SARS-CoV Severe acute respiratory syndrome coronavirus ASQAWQPGVAMPNLYKMQRMLLEKCDLQNYGENAVIPKGIMMNVAKYTQLCQYLNTLTLAVPYNMRVIHFGAGSDKGVAPGTAVLRQWLPTGTLLVDSDLNDFVSDADSTLIGDCATVHTANKWDLIISDMYDPRTKHVTKENDSKEGFFTYLCGFIKQKLALGGSIAVKITEHSWNADLYKLMGHFSWWTAFVTNVNASSSEAFLIGANYLGKPKEQIDGYTMHANYIFWRNTNPIQLSSYSLFDMSKFPLKLRGTAVMSLKENQINDMIYSLLEKGRLIIRENNRVVVSSDILVNN 6776 7073 14B6660399D3A801 s-adenosyl-l-methionine S adenosyl Methionine small molecule small molecule chemical synthesis Chemical synthesis (Chemical synthesis) p0c6x7-pro_0000037317 Non-structural protein 10 rep ORF1ab polyprotein 1a-1b protein protein sars-cov Human SARS coronavirus SARS-CoV Severe acute respiratory syndrome coronavirus AGNATEVPANSTVLSFCAFAVDPAKAYKDYLASGGQPITNCVKMLCTHTGTGQAITVTPEANMDQESFGGASCCLYCRCHIDHPNPKGFCDLKGKYVQIPTTCANDPVGFTLRNTVCTVCGMWKGYGCSCDQLREPLMQ 4231 4369 1F48DD3A6874826C rep-rep-90 1 7 unspecified role unspecified role bait bait over-expressed over expressed level over-expressed 9 unspecified role unspecified role prey prey over-expressed over expressed level over-expressed region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain physical association physical association 1 nsp10_nsp16_cvhsa-sars_rna_substrate-1 3 9 ancillary ancillary neutral component neutral component purified purified region required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain certain certain sequence position certain certain certain sequence position certain region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecoli Escherichia coli (strain K12) 7 enzyme enzyme neutral component neutral component purified purified n-terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range ecoli Escherichia coli (strain K12) 5 enzyme target enzyme target substrate substrate neutral component neutral component ATP 32p radiolabel 32p radiolabel P32 32P 32P P32 certain certain sequence position certain certain certain sequence position certain 2′-O-ribose methylated residue carbohydrate chemical modification carbohydrate chemical modification carbohydrate chemical modification certain certain sequence position certain certain certain sequence position certain resulting-ptm resulting-ptm resulting ptm resulting-ptm 5'_methyl_N7_Guanosine rna chemical modification RNA chemical modification RNA chemical modification post-transcriptional modification rna chemical modification certain certain sequence position certain certain certain sequence position certain in vitro In vitro methylation methylation reaction methylation 2C, Lanes 12,15,16. rep-rep-91 3 6 enzyme enzyme neutral component neutral component purified purified n-terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range ecoli Escherichia coli (strain K12) 7 enzyme enzyme neutral component neutral component purified purified n-terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range ecoli Escherichia coli (strain K12) 9 ancillary ancillary neutral component neutral component purified purified region required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain certain certain sequence position certain certain certain sequence position certain region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecoli Escherichia coli (strain K12) 5 enzyme target enzyme target substrate substrate neutral component neutral component 5'_methyl_N7_Guanosine rna chemical modification RNA chemical modification RNA chemical modification post-transcriptional modification rna chemical modification certain certain sequence position certain certain certain sequence position certain resulting-ptm resulting-ptm resulting ptm resulting-ptm 2′-O-ribose methylated residue carbohydrate chemical modification carbohydrate chemical modification carbohydrate chemical modification certain certain sequence position certain certain certain sequence position certain resulting-ptm resulting-ptm resulting ptm resulting-ptm ATP 32p radiolabel 32p radiolabel P32 32P 32P P32 certain certain sequence position certain certain certain sequence position certain 5'_Guanosine rna chemical modification RNA chemical modification RNA chemical modification post-transcriptional modification rna chemical modification certain certain sequence position certain certain certain sequence position certain in vitro In vitro methylation methylation reaction methylation 2C, Lanes 5,6,2. A two-staged reaction: overall reaction = nsp10/nsp14/nsp16 methylates G*pppA-RNA to m7G*pppAm-RNA (Fig. 2C, lane 2): nsp14 first methylates G*pppA-RNA at N7 position of guanine (m7G) to generate m7GpppA-RNA, (Fig. 2C, lanes 1 and 7), followed by the methylation of m7G*pppA-RNA at the 2′-O-position of adenine (Am) to generate m7G*pppAm-RNA. there are complexes for Nsp10 + nsp14, also Nsp10 + nsp16; but not appropriate to use for this reaction. rep-rep-92 3 6 enzyme enzyme neutral component neutral component purified purified n-terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range ecoli Escherichia coli (strain K12) 5 enzyme target enzyme target substrate substrate neutral component neutral component 5'_methyl_N7_Guanosine rna chemical modification RNA chemical modification RNA chemical modification post-transcriptional modification rna chemical modification certain certain sequence position certain certain certain sequence position certain resulting-ptm resulting-ptm resulting ptm resulting-ptm ATP 32p radiolabel 32p radiolabel P32 32P 32P P32 certain certain sequence position certain certain certain sequence position certain 5'_Guanosine rna chemical modification RNA chemical modification RNA chemical modification post-transcriptional modification rna chemical modification certain certain sequence position certain certain certain sequence position certain in vitro In vitro methylation methylation reaction methylation 2C, Lanes 1,7. Reaction: nsp14 methylates G*pppA-RNA at N7 position of guanine (m7G) to generate m7GpppA-RNA, (Fig. 2C, lanes 1 and 7). Either protein nsp10 or nsp16 was present but did not participate in the reaction. rep-rep-93 4 8 unspecified role unspecified role neutral component neutral component meres 3h radiolabel 3h radiolabel 3H tritium H3 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined chemical synthesis Chemical synthesis (Chemical synthesis) 9 stimulator stimulator neutral component neutral component purified purified region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecoli Escherichia coli (strain K12) 7 unspecified role unspecified role neutral component neutral component purified purified n-terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range ecoli Escherichia coli (strain K12) direct interaction direct interaction 2D, 4B All four truncated Nsp10 proteins attenuated the SAM binding affinity of nsp16. rep-rep-94 2 9 ancillary ancillary neutral component neutral component purified purified region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecoli Escherichia coli (strain K12) 7 enzyme enzyme neutral component neutral component purified purified n-terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range ecoli Escherichia coli (strain K12) 5 enzyme target enzyme target substrate substrate neutral component neutral component 2′-O-ribose methylated residue carbohydrate chemical modification carbohydrate chemical modification carbohydrate chemical modification certain certain sequence position certain certain certain sequence position certain resulting-ptm resulting-ptm resulting ptm resulting-ptm 5'_methyl_N7_Guanosine rna chemical modification RNA chemical modification RNA chemical modification post-transcriptional modification rna chemical modification certain certain sequence position certain certain certain sequence position certain in vitro In vitro methylation methylation reaction methylation 4A I did not use the complex CPX-5709 here, as I preferred to show construct details. rep-rep-95 2 9 competitor competitor competitor neutral component neutral component n-terminus ac1res monoacetylated residue Acetyl Acetylation (N terminus, N epsilon of Lysine, O of Serine) (Ac) Acetylation Ac1Res certain certain sequence position certain certain certain sequence position certain region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain C-terminus amidated residue amidated residue certain certain sequence position certain certain certain sequence position certain chemical synthesis Chemical synthesis (Chemical synthesis) 9 ancillary ancillary neutral component neutral component purified purified region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecoli Escherichia coli (strain K12) 7 enzyme enzyme neutral component neutral component purified purified n-terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range ecoli Escherichia coli (strain K12) 5 enzyme target enzyme target substrate substrate neutral component neutral component 5'_methyl_N7_Guanosine rna chemical modification RNA chemical modification RNA chemical modification post-transcriptional modification rna chemical modification certain certain sequence position certain certain certain sequence position certain 2′-O-ribose methylated residue carbohydrate chemical modification carbohydrate chemical modification carbohydrate chemical modification certain certain sequence position certain certain certain sequence position certain resulting-ptm resulting-ptm resulting ptm resulting-ptm in vitro In vitro methylation methylation reaction methylation 4A, I did not use the complex CPX-5709 here, as I preferred to show construct details. rep-rep-96 4 9 competitor competitor competitor neutral component neutral component n-terminus ac1res monoacetylated residue Acetyl Acetylation (N terminus, N epsilon of Lysine, O of Serine) (Ac) Acetylation Ac1Res certain certain sequence position certain certain certain sequence position certain region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain C-terminus amidated residue amidated residue certain certain sequence position certain certain certain sequence position certain chemical synthesis Chemical synthesis (Chemical synthesis) 8 unspecified role unspecified role neutral component neutral component meres 3h radiolabel 3h radiolabel 3H tritium H3 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined chemical synthesis Chemical synthesis (Chemical synthesis) 9 stimulator stimulator neutral component neutral component purified purified region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecoli Escherichia coli (strain K12) 7 unspecified role unspecified role neutral component neutral component purified purified n-terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range ecoli Escherichia coli (strain K12) direct interaction direct interaction 4B