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 Myosin filament assembly requires a cluster of four positive residues located in the rod domain. Myosin filament assembly requires a cluster of four positive residues located in the rod domain. FEBS Lett. (0014-5793) 2012 Thompson RC., Buvoli M., Buvoli A., Leinwand LA. suggested by lperfetto in vitro In vitro light scattering light scattering predetermined predetermined participant predetermined Thompson RC., Buvoli M., Buvoli A., Leinwand LA. suggested by lperfetto FEBS Lett. (0014-5793) 2012 Myosin filament assembly requires a cluster of four positive residues located in the rod domain. Myosin filament assembly requires a cluster of four positive residues located in the rod domain. FEBS Lett. (0014-5793) 2012 Thompson RC., Buvoli M., Buvoli A., Leinwand LA. suggested by lperfetto in vitro In vitro spr surface plasmon resonance Optical biosensor BIAcore(r) BIAcore(r) Optical biosensor spr predetermined predetermined participant predetermined Thompson RC., Buvoli M., Buvoli A., Leinwand LA. suggested by lperfetto FEBS Lett. (0014-5793) 2012 myh6_rat Myosin-6 Myh6 Myosin heavy chain 6 Myosin heavy chain, cardiac muscle alpha isoform protein protein rat Rattus norvegicus (Rat) Rat MTDAQMADFGAARYLRKSEKERLEAQTRPFDIRTECFVPDDKEEYVKAKIVSREGGKVTAETENGKTVTVKEDQVMQQNPPKFDKIEDMAMLTFLHEPAVLYNLKERYAAWMIYTYSGLFCVTVNPYKWLPVYNAEVVAAYRGKKRSEAPPHIFSISDNAYQYMLTDRENQSILITGESGAGKTVNTKRVIQYFASIAAIGDRSKKDNPNANKGTLEDQIIQANPALEAFGNAKTVRNDNSSRFGKFIRIHFGATGKLASADIETYLLEKSRVIFQLKAERNYHIFYQILSNKKPELLDMLLVTNNPYDYAFVSQGEVSVASIDDSEELLATDSAFDVLGFTAEEKAGVYKLTGAIMHYGNMKFKQKQREEQAEPDGTEDADKSAYLMGLNSADLLKGLCHPRVKVGNEYVTKGQSVQQVYYSIGALAKSVYEKMFNWMVTRINATLETKQPRQYFIGVLDIAGFEIFDFNSFEQLCINFTNEKLQQFFNHHMFVLEQEEYKKEGIEWEFIDFGMDLQACIDLIEKPMGIMSILEEECMFPKATDMTFKAKLYDNHLGKSNNFQKPRNVKGKQEAHFSLVHYAGTVDYNILGWLEKNKDPLNETVVGLYQKSSLKLMATLFSTYASADTGDSGKGKGGKKKGSSFQTVSALHRENLNKLMTNLRTTHPHFVRCIIPNERKAPGVMDNPLVMHQLRCNGVLEGIRICRKGFPNRILYGDFRQRYRILNPAAIPEGQFIDSGKGAEKLLGSLDIDHNQYKFGHTKVFFKAGLLGLLEEMRDERLSRIITRIQAQARGQLMRIEFKKMVERRDALLVIQWNIRAFMGVKNWPWMKLYFKIKPLLKSAETEKEMANMKEEFGRVKDALEKSEARRKELEEKMVSLLQEKNDLQLQVQAEQDNLADAEERCDQLIKNKIQLEAKVKEMTERLEDEEEMNAELTAKKRKLEDECSELKKDIDDLELTLAKVEKEKHATENKVKNLTEEMAGLDEIIAKLTKEKKALQEAHQQALDDLQAEEDKVNTLTKSKVKLEQQVDDLEGSLEQEKKVRMDLERAKRKLEGDLKLTQESIMDLENDKLQLEEKLKKKEFDISQQNSKIEDEQALALQLQKKLKENQARIEELEEELEAERTARAKVEKLRSDLTRELEEISERLEEAGGATSVQIEMNKKREAEFQKMRRDLEEATLQHEATAAALRKKHADSVAELGEQIDNLQRVKQKLEKEKSEFKLELDDVTSHMEQIIKAKANLEKVSRTLEDQANEYRVKLEEAQRSLNDFTTQRAKLQTENGELARQLEEKEALIWQLTRGKLSYTQQMEDLKRQLEEEGKAKNALAHALQSARHDCDLLREQYEEEMEAKAELQRVLSKANSEVAQWRTKYETDAIQRTEELEEAKKKLAQRLQDAEEAVEAVNAKCSSLEKTKHRLQNEIEDLMVDVERSNAAAAALDKKQRNFDKILAEWKQKYEESQSELESSQKEARSLSTELFKLKNAYEESLEHLETFKRENKNLQEEISDLTEQLGEGGKNVHELEKIRKQLEVEKLELQSALEEAEASLEHEEGKILRAQLEFNQIKAEIERKLAEKDEEMEQAKRNHLRVVDSLQTSLDAETRSRNEALRVKKKMEGDLNEMEIQLSQANRIASEAQKHLKNAQAHLKDTQLQLDDAVRANDDLKENIAIVERRNTLLQAELEELRAVVEQTERSRKLAEQELIETSERVQLLHSQNTSLINQKKKMDADLSQLQTEVEEAVQECRNAEEKAKKAITDAAMMAEELKKEQDTSAHLERMKKNMEQTIKDLQHRLDEAEQIALKGGKKQLQKLEARVRELENELEAEQKRNAESVKGMRKSERRIKELTYQTEEDKKNLVRLQDLVDKLQLKVKAYKRQAEEAEEQANTNLSKFRKVQHELDEAEERADIAESQVNKLRAKSRDIGAKQKMHDEE DIP protein P02563 original sequence version: 126 D7BD33FC2B19E3C2 myh6-myh6 2 WT LMM 3 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role bait bait purified purified binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain tandem tag tandem tag tandem tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined WT LMM 3 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role prey prey purified purified tandem tag tandem tag tandem tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain direct interaction direct interaction f1 homomint mint In our assay, we measure the noncovalent binding of LMM analyte in solution to the LMM ligand immobilized on the biosensor chip. As the ligand and analyte interact, the formation of a myosin polymer on the chip increases the refractive index of the solution proportionally to the mass of analyte bound. Figure 1A shows the results of a typical experiment in which WT-LMM ligand (residues 1231-1938), covalently attached to the chip through a Cys at its N-terminus, was exposed to solutions of WT-LMM analyte in 150 mM NaCl at pH 7.3. At analyte concentrations greater than 0.2 μM the amount of LMM bound increases with time with a rate that is approximately first order in protein concentration. Since the amount of noncovalently bound LMM analyte quickly exceeds stoichiometric equivalence with the covalently bound LMM ligand (see Materials and Methods) the LMM analyte is either polymerizing on the LMM ligand, or is polymerizing in solution and then binding to the LMM ligand. The complex formed under these conditions dissociates rapidly when highsalt buffer replaces the analyte (asterisks in each panels). The kinetics of dissociation are complex and do not lend themselves to a simple analysis of this process. myh6-myh6-1 2 WT LMM 3 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role bait bait purified purified binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain tandem tag tandem tag tandem tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined alanine mutant LMM 3 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role prey prey purified purified binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain P02563:p.Arg1922Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain R A Tue Jul 12 10:49:09 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 14:43:14 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'arg1922ala' Tue Jun 30 19:36:16 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Arg1922Ala' P02563:p.Lys1920Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain K A Tue Jul 12 10:49:20 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 16:23:09 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys1920ala' Tue Jun 30 16:45:19 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Lys1920Ala' P02563:p.Lys1924Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain K A Tue Jul 12 10:06:12 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 18:44:53 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys1924ala' Tue Jun 30 17:44:03 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Lys1924Ala' tandem tag tandem tag tandem tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined P02563:p.Arg1926Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain R A Tue Jul 12 10:06:15 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 15:08:10 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'arg1926ala' Tue Jun 30 19:45:31 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Arg1926Ala' direct interaction direct interaction f2B homomint mint To further localize the residues in the C-terminal region identified by the truncation analysis as being important for the polymerization reaction, we closely examined the sequence contained in the CT2 deletion and focused on a cluster of four charged amino acids, (KLRAKSR, residues 1920-1926) which we postulated could be involved in ionic interactions between adjacent coiled-coils since present in the exposed positions of the heptad repeat (Figure 2, bottom). In accord with our hypothesis, simultaneous replacement of all four positively charged residues with alanine reduced the LMM analyte s ability to bind to the LMM ligand (Figure 2B). Interestingly, the alanine replacement mutagenesis has a greater impact on polymerization than the CT2 truncation. myh6-myh6-2 2 CT2 LMM 3 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role prey prey purified purified undefined mutation mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain Wed Jul 06 16:27:55 BST 2016 Sequence change details about this feature cannot be ascertained or do not fit with the current version of the referenced protein, so they have been deleted as a result of our quality control procedures. The original label was 'mutation disrupting' 1919-1938 tandem tag tandem tag tandem tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain WT LMM 3 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role bait bait purified purified tandem tag tandem tag tandem tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain direct interaction direct interaction homomint mint f2A To further define those amino acid 6 residues responsible for assembly we have investigated the properties of a number of C-terminally truncated LMM analytes in the SPR assay (Figure 2A). While binding was hardly affected by truncation of the C-terminal 6 amino acids (CT1), it was severely reduced by truncation of the C-terminal 20 residues of LMM (CT2). myh6-myh6-3 1 alanine mutant LMM 3 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role neutral component neutral component purified purified P02563:p.Lys1920Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain K A Tue Jul 12 10:50:14 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 18:03:36 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys1920ala' Tue Jun 30 15:40:07 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Lys1920Ala' tandem tag tandem tag tandem tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined P02563:p.Arg1922Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain R A Tue Jul 12 10:50:09 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' 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 'arg1922ala' Tue Jun 30 18:08:05 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Arg1922Ala' P02563:p.Arg1926Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain R A Tue Jul 12 10:50:10 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 16:52:31 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'arg1926ala' Tue Jun 30 19:10:49 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Arg1926Ala' binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain P02563:p.Lys1924Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain K A Tue Jul 12 10:50:24 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 19:11:01 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys1924ala' Tue Jun 30 16:14:43 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Lys1924Ala' alanine mutant LMM 3 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role neutral component neutral component purified purified P02563:p.Lys1920Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain K A Tue Jul 12 10:07:41 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 16:44:31 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys1920ala' Tue Jun 30 18:40:28 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Lys1920Ala' P02563:p.Arg1922Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain R A Tue Jul 12 10:07:45 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 17:34:07 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'arg1922ala' Tue Jun 30 17:00:55 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Arg1922Ala' tandem tag tandem tag tandem tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain P02563:p.Lys1924Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain K A Tue Jul 12 10:07:37 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 20:30:53 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys1924ala' Tue Jun 30 20:12:21 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Lys1924Ala' P02563:p.Arg1926Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain R A Tue Jul 12 10:08:01 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 18:07:58 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'arg1926ala' Tue Jun 30 17:18:35 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Arg1926Ala' direct interaction direct interaction homomint mint f3B The absence of any measurable increase in light scattering observed when the C-terminal KLRAKSR/ALAAASA mutant was assayed indicates that the loss of the four positive residues severely affects the ability of LMM to self-polymerize (Figure 3B, red curve). This result also suggests that the dramatic signal reduction observed in the SPR assay with the same mutant is probably due to lack of analyte self-polymerization. myh6-myh6-4 1 alanine mutant LMM 3 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role neutral component neutral component purified purified binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain P02563:p.Arg1922Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain R A Tue Jul 12 10:07:52 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 16:05:13 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'arg1922ala' Tue Jun 30 19:08:38 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Arg1922Ala' P02563:p.Arg1926Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain R A Tue Jul 12 10:07:53 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 16:02:50 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'arg1926ala' Tue Jun 30 17:49:14 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Arg1926Ala' P02563:p.Lys1924Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain K A Tue Jul 12 10:50:21 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 20:24:24 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys1924ala' Tue Jun 30 18:24:50 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Lys1924Ala' tandem tag tandem tag tandem tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined P02563:p.Lys1920Ala mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain K A Tue Jul 12 10:08:12 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 15:06:36 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys1920ala' Tue Jun 30 19:45:06 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Lys1920Ala' WT LMM 3 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role neutral component neutral component purified purified binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain tandem tag tandem tag tandem tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined direct interaction direct interaction homomint mint f3B Moreover, mixing equimolar amounts of the ALAAASA mutant with wild-type LMM only slightly enhanced the extent of light scattering (Figure 3B, purple curve) indicating that the mutant cannot efficiently copolymerize with, or inhibit the polymerization of the wild-type protein myh6-myh6-5 1 WT LMM 3 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role neutral component neutral component purified purified tandem tag tandem tag tandem tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain WT LMM 3 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role neutral component neutral component purified purified tandem tag tandem tag tandem tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain direct interaction direct interaction homomint mint f3A As for SPR, the LS assay shows that LMM polymerization is dependent on its concentration (Figure 3A) and on the ionic strength of the solution (data not shown).