MINT MINT, Dpt of Biology, University of Rome Tor Vergata MINT http://mint.bio.uniroma2.it/mint url>http://mint.bio.uniroma2.it/mint/ https://mint.bio.uniroma2.it MINT-[0-9]+ https://mint.bio.uniroma2.it/index.php/results-interactions/?id=${ac} The Molecular INTeraction database (MINT) is a relational database designed to store interactions between biological molecules. http://mint.bio.uniroma2.it Identification of amino acid residues in HIV-1 reverse transcriptase that are critical for the proteolytic processing of Gag-Pol precursors. Identification of amino acid residues in HIV-1 reverse transcriptase that are critical for the proteolytic processing of Gag-Pol precursors. FEBS Lett. (0014-5793) 2011 imex curation Nishitsuji H., Yokoyama M., Sato H., Yamauchi S., Takaku H. suggested by lperfetto hironori.nishitsuji@it-chiba.ac.jp Only protein-protein interactions in vitro In vitro protein kinase assay protein kinase assay western blot western blot Immuno blot Immuno blot Nishitsuji H., Yokoyama M., Sato H., Yamauchi S., Takaku H. suggested by lperfetto hironori.nishitsuji@it-chiba.ac.jp FEBS Lett. (0014-5793) 2011 Only protein-protein interactions imex curation Identification of amino acid residues in HIV-1 reverse transcriptase that are critical for the proteolytic processing of Gag-Pol precursors. Identification of amino acid residues in HIV-1 reverse transcriptase that are critical for the proteolytic processing of Gag-Pol precursors. FEBS Lett. (0014-5793) 2011 imex curation Nishitsuji H., Yokoyama M., Sato H., Yamauchi S., Takaku H. suggested by lperfetto hironori.nishitsuji@it-chiba.ac.jp Only protein-protein interactions human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. crosslink cross-linking study crosslink western blot western blot Immuno blot Immuno blot Nishitsuji H., Yokoyama M., Sato H., Yamauchi S., Takaku H. suggested by lperfetto hironori.nishitsuji@it-chiba.ac.jp FEBS Lett. (0014-5793) 2011 Only protein-protein interactions imex curation csk21_human Casein kinase II subunit alpha CSNK2A1 CK2A1 protein protein human Homo sapiens Human MSGPVPSRARVYTDVNTHRPREYWDYESHVVEWGNQDDYQLVRKLGRGKYSEVFEAINITNNEKVVVKILKPVKKKKIKREIKILENLRGGPNIITLADIVKDPVSRTPALVFEHVNNTDFKQLYQTLTDYDIRFYMYEILKALDYCHSMGIMHRDVKPHNVMIDHEHRKLRLIDWGLAEFYHPGQEYNVRVASRYFKGPELLVDYQMYDYSLDMWSLGCMLASMIFRKEPFFHGHDNYDQLVRIAKVLGTEDLYDYIDKYNIELDPRFNDILGRHSRKRWERFVHSENQHLVSPEALDFLDKLLRYDHQSRLTAREAMEHPYFYTVVKDQARMGSSSMPGGSTPVSSANMMSGISSVPTPSPLGPLAGSPVIAAANPLGMPVPAAAGAQQ DIP protein P68400 original sequence version: 88 DIP protein P68400 original sequence version: 78 DIP protein P68400 original sequence version: 100 DIP protein P68400 original sequence version: 110 DIP protein P68400 original sequence version: 125 DIP protein P68400 original sequence version: 136 D3B6F5D13FF7422D q72547_9hiv1 pol protein protein 9hiv1 Human immunodeficiency virus 1 PISPIETVPVKLKPGMDGPKVKQWPLTEEKIKALVEICTEMEKEGKISKIGPENPYNTPVFAIKKKDSTKWRKLVDFRELNKRTQDFWEVQLGIPHPAGLKKRKSVTVLDVGDAYFSVPLDEDFRKYTAFTIPSINNETPGIRYQYNVLPQGWKGSPAIFQSSMTKILEPFRKQNPDIVIYQYMDDLYVGSDLEIGQHRTKIEELRQHLLRWGLTTPDKKHQKEPPFLWMGYELHPDKWTVQPIVLPEKDSWTVNDIQKLVGKLNWASQIYPGIRVRQLCKLLRGTKALTEVIPLTEEAELELAENREILKEPVHGVYYDPSKDLIAEIQKQGQGQWTYQIYQEPFKNLRTGKYARMRGAHTNDVKQLTEAVQKITTESIVIWGKTPKFKLPIQKETWETWWTEYWQATWIPEWEFVNTPPLVKLWYQLEKEPIVGAETFYVDGAANRETKLGKAGYVTNRGRQKVVTLTDTTNQKTELQAIYLALQDSGLEVNIVTDSQYALGIIQAQPDQSESELVNQIIEQLIKKEKVYLAWVPAHKGIGGNEQVDKLVSAGIRKVLFLDGID mint AA4B9287ECC4E5C6 pol-2 2 RT 4 western blot western blot Immuno blot Immuno blot predetermined predetermined participant predetermined unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed living cell living cell binding site binding region binding-associated region binding region undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined - [EBI-7484791]Certain and ragged-n-terminus positions must always be strictly superior to 0 and inferior or equal to the protein sequence length. Actual positions : 600-600, sequence length 566 [EBI-7484791]600-1159 rt RT polyprotein frag polyprotein fragment polyprotein frag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined [EBI-7484796]Certain and ragged-n-terminus positions must always be strictly superior to 0 and inferior or equal to the protein sequence length. Actual positions : 600-600, sequence length 566 [EBI-7484796]600-1159 v5 tag v5 tag v5 tag GKPIPNPLLGLDST epitope tag GKPIPNPLLGLDST epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined Q72547:p.Thr131Ala mutation decreasing interaction mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain T A Tue Jul 12 10:58:30 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation decreasing' Wed May 02 15:51:45 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr131ala' Tue Jun 30 19:48:25 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr131Ala' Stoichiometry: 2.0 physical association physical association f3 mint The addition of 0.2 mM DTSSP yielded cross-linked complexes of RT homodimers when RT-WT-V5 was expressed in 293T cells (Fig. 3). A T131A mutation slightly reduced the level of RT homodimers in the presence of DTSSP compared with the wild type. In contrast, an abnormal pattern was observed in the Y146A mutant in the absence or presence of DTSSP, suggesting that Y146A induced improper folding. Moreover, a T128A mutation resulted in very low levels of the dimer under the same experimental conditions. T58A, Y183A and T216A mutations, which resulted in normal Gag-Pol, retained RT dimerization in the presence of DTSSP. These results suggest that RT dimerization may affect Gag-Pol processing. However, the possibility that the T128A andY146A mutations significantly change the conformation of Gag-Pol precursor, decreasing the accessibility of the targets, cannot be excluded. pol-2 2 RT 4 western blot western blot Immuno blot Immuno blot predetermined predetermined participant predetermined unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed living cell living cell v5 tag v5 tag v5 tag GKPIPNPLLGLDST epitope tag GKPIPNPLLGLDST epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined Q72547:p.Tyr146Ala mutation decreasing interaction mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain Y A Tue Jul 12 10:06:02 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation decreasing' Wed May 02 14:50:54 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'tyr146ala' Tue Jun 30 16:18:28 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Tyr146Ala' rt RT polyprotein frag polyprotein fragment polyprotein frag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined [EBI-7484936]Certain and ragged-n-terminus positions must always be strictly superior to 0 and inferior or equal to the protein sequence length. Actual positions : 600-600, sequence length 566 [EBI-7484936]600-1159 binding site binding region binding-associated region binding region undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined - [EBI-7484942]Certain and ragged-n-terminus positions must always be strictly superior to 0 and inferior or equal to the protein sequence length. Actual positions : 600-600, sequence length 566 [EBI-7484942]600-1159 Stoichiometry: 2.0 physical association physical association f3 mint The addition of 0.2 mM DTSSP yielded cross-linked complexes of RT homodimers when RT-WT-V5 was expressed in 293T cells (Fig. 3). A T131A mutation slightly reduced the level of RT homodimers in the presence of DTSSP compared with the wild type. In contrast, an abnormal pattern was observed in the Y146A mutant in the absence or presence of DTSSP, suggesting that Y146A induced improper folding. Moreover, a T128A mutation resulted in very low levels of the dimer under the same experimental conditions. T58A, Y183A and T216A mutations, which resulted in normal Gag-Pol, retained RT dimerization in the presence of DTSSP. These results suggest that RT dimerization may affect Gag-Pol processing. However, the possibility that the T128A andY146A mutations significantly change the conformation of Gag-Pol precursor, decreasing the accessibility of the targets, cannot be excluded. pol-2 2 RT 4 western blot western blot Immuno blot Immuno blot predetermined predetermined participant predetermined unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed living cell living cell Q72547:p.Thr128Ala mutation decreasing interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain T A Tue Jul 12 10:06:16 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation decreasing' Wed May 02 14:41:31 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr128ala' Tue Jun 30 16:13:17 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr128Ala' rt RT polyprotein frag polyprotein fragment polyprotein frag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined [EBI-7484982]Certain and ragged-n-terminus positions must always be strictly superior to 0 and inferior or equal to the protein sequence length. Actual positions : 600-600, sequence length 566 [EBI-7484982]600-1159 v5 tag v5 tag v5 tag GKPIPNPLLGLDST epitope tag GKPIPNPLLGLDST epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined Stoichiometry: 2.0 physical association physical association f3 mint The addition of 0.2 mM DTSSP yielded cross-linked complexes of RT homodimers when RT-WT-V5 was expressed in 293T cells (Fig. 3). A T131A mutation slightly reduced the level of RT homodimers in the presence of DTSSP compared with the wild type. In contrast, an abnormal pattern was observed in the Y146A mutant in the absence or presence of DTSSP, suggesting that Y146A induced improper folding. Moreover, a T128A mutation resulted in very low levels of the dimer under the same experimental conditions. T58A, Y183A and T216A mutations, which resulted in normal Gag-Pol, retained RT dimerization in the presence of DTSSP. These results suggest that RT dimerization may affect Gag-Pol processing. However, the possibility that the T128A andY146A mutations significantly change the conformation of Gag-Pol precursor, decreasing the accessibility of the targets, cannot be excluded. csnk2a1-pol 1 RT 4 western blot western blot Immuno blot Immuno blot predetermined predetermined participant predetermined enzyme target enzyme target substrate substrate neutral component neutral component purified purified his tag 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 phosphorylated phosphorylated residue phosres phosphorylated residue PhosRes Phospho Phosphorylation Phosphorylation (O of Serine, Threonine, Tyrosine and Aspartate, N epsilon of Lysine) mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain resulting-ptm phosphorylated phosphorylated residue phosres phosphorylated residue PhosRes Phospho Phosphorylation Phosphorylation (O of Serine, Threonine, Tyrosine and Aspartate, N epsilon of Lysine) mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain resulting-ptm rt RT polyprotein frag polyprotein fragment polyprotein frag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined [EBI-7485059]Certain and ragged-n-terminus positions must always be strictly superior to 0 and inferior or equal to the protein sequence length. Actual positions : 600-600, sequence length 566 [EBI-7485059]600-1159 binding site binding region binding-associated region binding region undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined - [EBI-7485065]Certain and ragged-n-terminus positions must always be strictly superior to 0 and inferior or equal to the protein sequence length. Actual positions : 600-600, sequence length 566 [EBI-7485065]600-1159 CK2 alpha 3 western blot western blot Immuno blot Immuno blot predetermined predetermined participant predetermined enzyme enzyme neutral component neutral component purified purified phosphorylation phosphorylation reaction phosphorylation homomint mint f4a f4b f4c As shown in Figure 4A, a shifted band of recombinant His-RT-WT was observed in the presence of recombinant CK2-alpha in a dose-dependent manner. We next examined the phosphorylation of mutant RT. The phosphorylation levels of His-RT-T128A were similar to wild-type RT (Fig. 4B). In the case of the His-RT-Y146A mutants, the phosphorylation levels were slightly reduced compared with wild-type levels. Moreover, both the T128A and Y146A mutations in recombinant His-RT severely decreased the phosphorylation levels. However, a shifted band corresponding to phosphorylated His-RT-T128A-Y146A could be detected by using a large amount of recombinant His-RT-T128A-Y146A in an in vitro kinase assay (Fig. 4C)