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Yorodumi- EMDB-28270: Helical reconstruction of the human cardiac actin-tropomyosin-myo... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-28270 | |||||||||
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Title | Helical reconstruction of the human cardiac actin-tropomyosin-myosin loop 4 7G mutant complex | |||||||||
Map data | Primary map used for model building. | |||||||||
Sample |
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Keywords | actin / tropomyosin / myosin / cardiac / MOTOR PROTEIN | |||||||||
Function / homology | Function and homology information positive regulation of heart rate by epinephrine / muscle thin filament tropomyosin / regulation of slow-twitch skeletal muscle fiber contraction / regulation of the force of skeletal muscle contraction / actin-myosin filament sliding / bleb / negative regulation of vascular associated smooth muscle cell migration / regulation of muscle contraction / muscle myosin complex / muscle filament sliding ...positive regulation of heart rate by epinephrine / muscle thin filament tropomyosin / regulation of slow-twitch skeletal muscle fiber contraction / regulation of the force of skeletal muscle contraction / actin-myosin filament sliding / bleb / negative regulation of vascular associated smooth muscle cell migration / regulation of muscle contraction / muscle myosin complex / muscle filament sliding / transition between fast and slow fiber / regulation of the force of heart contraction / myosin filament / adult heart development / ruffle organization / cardiac muscle hypertrophy in response to stress / negative regulation of vascular associated smooth muscle cell proliferation / positive regulation of ATP-dependent activity / Striated Muscle Contraction / myosin II complex / myosin complex / structural constituent of muscle / ventricular cardiac muscle tissue morphogenesis / regulation of heart contraction / sarcomere organization / microfilament motor activity / myosin binding / myofibril / heart contraction / mesenchyme migration / skeletal muscle contraction / positive regulation of cell adhesion / Smooth Muscle Contraction / striated muscle contraction / muscle contraction / cardiac muscle contraction / stress fiber / ATP metabolic process / positive regulation of stress fiber assembly / cytoskeleton organization / cytoskeletal protein binding / regulation of heart rate / sarcomere / negative regulation of cell migration / filopodium / actin filament organization / actin filament / ruffle membrane / wound healing / cellular response to reactive oxygen species / structural constituent of cytoskeleton / Z disc / actin filament binding / actin cytoskeleton / lamellipodium / actin binding / regulation of cell shape / cell body / calmodulin binding / cytoskeleton / protein heterodimerization activity / positive regulation of gene expression / protein homodimerization activity / ATP binding / identical protein binding / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Sus scrofa (pig) / Homo sapiens (human) | |||||||||
Method | helical reconstruction / cryo EM / Resolution: 3.6 Å | |||||||||
Authors | Doran MH / Lehman W / Rynkiewicz MJ | |||||||||
Funding support | United States, 1 items
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Citation | Journal: J Gen Physiol / Year: 2023 Title: Myosin loop-4 is critical for optimal tropomyosin repositioning on actin during muscle activation and relaxation. Authors: Matthew H Doran / Michael J Rynkiewicz / Elumalai Pavadai / Skylar M L Bodt / David Rasicci / Jeffrey R Moore / Christopher M Yengo / Esther Bullitt / William Lehman / Abstract: During force-generating steps of the muscle crossbridge cycle, the tip of the myosin motor, specifically loop-4, contacts the tropomyosin cable of actin filaments. In the current study, we determined ...During force-generating steps of the muscle crossbridge cycle, the tip of the myosin motor, specifically loop-4, contacts the tropomyosin cable of actin filaments. In the current study, we determined the corresponding effect of myosin loop-4 on the regulatory positioning of tropomyosin on actin. To accomplish this, we compared high-resolution cryo-EM structures of myosin S1-decorated thin filaments containing either wild-type or a loop-4 mutant construct, where the seven-residue portion of myosin loop-4 that contacts tropomyosin was replaced by glycine residues, thus removing polar side chains from residues 366-372. Cryo-EM analysis of fully decorated actin-tropomyosin filaments with wild-type and mutant S1, yielded 3.4-3.6 Å resolution reconstructions, with even higher definition at the actin-myosin interface. Loop-4 densities both in wild-type and mutant S1 were clearly identified, and side chains were resolved in the wild-type structure. Aside from loop-4, actin and myosin structural domains were indistinguishable from each other when filaments were decorated with either mutant or wild-type S1. In marked contrast, the position of tropomyosin on actin in the two reconstructions differed by 3 to 4 Å. In maps of filaments containing the mutant, tropomyosin was located closer to the myosin-head and thus moved in the direction of the C-state conformation adopted by myosin-free thin filaments. Complementary interaction energy measurements showed that tropomyosin in the mutant thin filaments sits on actin in a local energy minimum, whereas tropomyosin is positioned by wild-type S1 in an energetically unfavorable location. We propose that the high potential energy associated with tropomyosin positioning in wild-type filaments favors an effective transition to B- and C-states following release of myosin from the thin filaments during relaxation. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_28270.map.gz | 233.6 MB | EMDB map data format | |
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Header (meta data) | emd-28270-v30.xml emd-28270.xml | 25.4 KB 25.4 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_28270_fsc.xml | 17 KB | Display | FSC data file |
Images | emd_28270.png | 61.5 KB | ||
Masks | emd_28270_msk_1.map | 421.9 MB | Mask map | |
Filedesc metadata | emd-28270.cif.gz | 8.3 KB | ||
Others | emd_28270_additional_1.map.gz emd_28270_half_map_1.map.gz emd_28270_half_map_2.map.gz | 390.2 MB 337.2 MB 337.1 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-28270 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-28270 | HTTPS FTP |
-Validation report
Summary document | emd_28270_validation.pdf.gz | 892.3 KB | Display | EMDB validaton report |
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Full document | emd_28270_full_validation.pdf.gz | 891.9 KB | Display | |
Data in XML | emd_28270_validation.xml.gz | 24.8 KB | Display | |
Data in CIF | emd_28270_validation.cif.gz | 33 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-28270 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-28270 | HTTPS FTP |
-Related structure data
Related structure data | 8encMC 8efiC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_28270.map.gz / Format: CCP4 / Size: 421.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | Primary map used for model building. | ||||||||||||||||||||
Voxel size | X=Y=Z: 1.078 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_28270_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: Uses the same data as the primary map,...
File | emd_28270_additional_1.map | ||||||||||||
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Annotation | Uses the same data as the primary map, but is globally filtered from the relion postprocessing job. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map 1.
File | emd_28270_half_map_1.map | ||||||||||||
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Annotation | Half map 1. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map 2.
File | emd_28270_half_map_2.map | ||||||||||||
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Annotation | Half map 2. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Human cardiac actin-tropomyosin-beta-myosin II complex with 7G lo...
Entire | Name: Human cardiac actin-tropomyosin-beta-myosin II complex with 7G loop 4 mutation |
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Components |
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-Supramolecule #1: Human cardiac actin-tropomyosin-beta-myosin II complex with 7G lo...
Supramolecule | Name: Human cardiac actin-tropomyosin-beta-myosin II complex with 7G loop 4 mutation type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3 Details: Cardiac actomyosin-tropomyosin complex with seven loop 4 residues mutated to glycine. |
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-Supramolecule #2: Cardiac F-actin complex
Supramolecule | Name: Cardiac F-actin complex / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #2 / Details: F-actin forms the backbone of the complex |
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Source (natural) | Organism: Sus scrofa (pig) |
-Supramolecule #3: Human beta-cardiac myosin II with seven (366-372) loop 4 residues...
Supramolecule | Name: Human beta-cardiac myosin II with seven (366-372) loop 4 residues mutated to glycine. type: complex / ID: 3 / Parent: 1 / Macromolecule list: #1 Details: The motor domain of the myosin saturates the actin filament. |
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Source (natural) | Organism: Homo sapiens (human) |
-Supramolecule #4: Human cardiac tropomyosin
Supramolecule | Name: Human cardiac tropomyosin / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #3 / Details: Tropomyosin wraps around the F-actin core. |
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Source (natural) | Organism: Homo sapiens (human) |
-Macromolecule #1: Myosin-7
Macromolecule | Name: Myosin-7 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 222.931328 KDa |
Recombinant expression | Organism: Mus musculus (house mouse) |
Sequence | String: MGDSEMAVFG AAAPYLRKSE KERLEAQTRP FDLKKDVFVP DDKQEFVKAK IVSREGGKVT AETEYGKTVT VKEDQVMQQN PPKFDKIED MAMLTFLHEP AVLYNLKDRY GSWMIYTYSG LFCVTVNPYK WLPVYTPEVV AAYRGKKRSE APPHIFSISD N AYQYMLTD ...String: MGDSEMAVFG AAAPYLRKSE KERLEAQTRP FDLKKDVFVP DDKQEFVKAK IVSREGGKVT AETEYGKTVT VKEDQVMQQN PPKFDKIED MAMLTFLHEP AVLYNLKDRY GSWMIYTYSG LFCVTVNPYK WLPVYTPEVV AAYRGKKRSE APPHIFSISD N AYQYMLTD RENQSILITG ESGAGKTVNT KRVIQYFAVI AAIGDRSKKD QSPGKGTLED QIIQANPALE AFGNAKTVRN DN SSRFGKF IRIHFGATGK LASADIETYL LEKSRVIFQL KAERDYHIFY QILSNKKPEL LDMLLITNNP YDYAFISQGE TTV ASIDDA EELMATDNAF DVLGFTSEEK NSMYKLTGAI MHFGNMKFKG GGGGGGAEPD GTEEADKSAY LMGLNSADLL KGLC HPRVK VGNEYVTKGQ NVQQVIYATG ALAKAVYERM FNWMVTRINA TLETKQPRQY FIGVLDIAGF EIFDFNSFEQ LCINF TNEK LQQFFNHHMF VLEQEEYKKE GIEWTFIDFG MDLQACIDLI EKPMGIMSIL EEECMFPKAT DMTFKAKLFD NHLGKS ANF QKPRNIKGKP EAHFSLIHYA GIVDYNIIGW LQKNKDPLNE TVVGLYQKSS LKLLSTLFAN YAGADAPIEK GKGKAKK GS SFQTVSALHR ENLNKLMTNL RSTHPHFVRC IIPNETKSPG VMDNPLVMHQ LRCNGVLEGI RICRKGFPNR ILYGDFRQ R YRILNPAAIP EGQFIDSRKG AEKLLSSLDI DHNQYKFGHT KVFFKAGLLG LLEEMRDERL SRIITRIQAQ SRGVLARME YKKLLERRDS LLVIQWNIRA FMGVKNWPWM KLYFKIKPLL KSAEREKEMA SMKEEFTRLK EALEKSEARR KELEEKMVSL LQEKNDLQL QVQAEQDNLA DAEERCDQLI KNKIQLEAKV KEMNERLEDE EEMNAELTAK KRKLEDECSE LKRDIDDLEL T LAKVEKEK HATENKVKNL TEEMAGLDEI IAKLTKEKKA LQEAHQQALD DLQAEEDKVN TLTKAKVKLE QQVDDLEGSL EQ EKKVRMD LERAKRKLEG DLKLTQESIM DLENDKQQLD ERLKKKDFEL NALNARIEDE QALGSQLQKK LKELQARIEE LEE ELEAER TARAKVEKLR SDLSRELEEI SERLEEAGGA TSVQIEMNKK REAEFQKMRR DLEEATLQHE ATAAALRKKH ADSV AELGE QIDNLQRVKQ KLEKEKSEFK LELDDVTSNM EQIIKAKANL EKMCRTLEDQ MNEHRSKAEE TQRSVNDLTS QRAKL QTEN GELSRQLDEK EALISQLTRG KLTYTQQLED LKRQLEEEVK AKNALAHALQ SARHDCDLLR EQYEEETEAK AELQRV LSK ANSEVAQWRT KYETDAIQRT EELEEAKKKL AQRLQEAEEA VEAVNAKCSS LEKTKHRLQN EIEDLMVDVE RSNAAAA AL DKKQRNFDKI LAEWKQKYEE SQSELESSQK EARSLSTELF KLKNAYEESL EHLETFKREN KNLQEEISDL TEQLGSSG K TIHELEKVRK QLEAEKMELQ SALEEAEASL EHEEGKILRA QLEFNQIKAE IERKLAEKDE EMEQAKRNHL RVVDSLQTS LDAETRSRNE ALRVKKKMEG DLNEMEIQLS HANRMAAEAQ KQVKSLQSLL KDTQIQLDDA VRANDDLKEN IAIVERRNNL LQAELEELR AVVEQTERSR KLAEQELIET SERVQLLHSQ NTSLINQKKK MDADLSQLQT EVEEAVQECR NAEEKAKKAI T DAAMMAEE LKKEQDTSAH LERMKKNMEQ TIKDLQHRLD EAEQIALKGG KKQLQKLEAR VRELENELEA EQKRNAESVK GM RKSERRI KELTYQTEED RKNLLRLQDL VDKLQLKVKA YKRQAEEAEE QANTNLSKFR KVQHELDEAE ERADIAESQV NKL RAKSRD IGTKGLNEE UniProtKB: Myosin-7 |
-Macromolecule #2: Actin, alpha cardiac muscle 1
Macromolecule | Name: Actin, alpha cardiac muscle 1 / type: protein_or_peptide / ID: 2 / Number of copies: 5 / Enantiomer: LEVO |
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Source (natural) | Organism: Sus scrofa (pig) |
Molecular weight | Theoretical: 42.064891 KDa |
Sequence | String: MCDDEETTAL VCDNGSGLVK AGFAGDDAPR AVFPSIVGRP RHQGVMVGMG QKDSYVGDEA QSKRGILTLK YPIEHGIITN WDDMEKIWH HTFYNELRVA PEEHPTLLTE APLNPKANRE KMTQIMFETF NVPAMYVAIQ AVLSLYASGR TTGIVLDSGD G VTHNVPIY ...String: MCDDEETTAL VCDNGSGLVK AGFAGDDAPR AVFPSIVGRP RHQGVMVGMG QKDSYVGDEA QSKRGILTLK YPIEHGIITN WDDMEKIWH HTFYNELRVA PEEHPTLLTE APLNPKANRE KMTQIMFETF NVPAMYVAIQ AVLSLYASGR TTGIVLDSGD G VTHNVPIY EGYALPHAIM RLDLAGRDLT DYLMKILTER GYSFVTTAER EIVRDIKEKL CYVALDFENE MATAASSSSL EK SYELPDG QVITIGNERF RCPETLFQPS FIGMESAGIH ETTYNSIMKC DIDIRKDLYA NNVLSGGTTM YPGIADRMQK EIT ALAPST MKIKIIAPPE RKYSVWIGGS ILASLSTFQQ MWISKQEYDE AGPSIVHRKC F UniProtKB: Actin, alpha cardiac muscle 1 |
-Macromolecule #3: Tropomyosin alpha-1 chain
Macromolecule | Name: Tropomyosin alpha-1 chain / type: protein_or_peptide / ID: 3 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 32.763621 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: MDAIKKKMQM LKLDKENALD RAEQAEADKK AAEDRSKQLE DELVSLQKKL KGTEDELDKY SEALKDAQEK LELAEKKATD AEADVASLN RRIQLVEEEL DRAQERLATA LQKLEEAEKA ADESERGMKV IESRAQKDEE KMEIQEIQLK EAKHIAEDAD R KYEEVARK ...String: MDAIKKKMQM LKLDKENALD RAEQAEADKK AAEDRSKQLE DELVSLQKKL KGTEDELDKY SEALKDAQEK LELAEKKATD AEADVASLN RRIQLVEEEL DRAQERLATA LQKLEEAEKA ADESERGMKV IESRAQKDEE KMEIQEIQLK EAKHIAEDAD R KYEEVARK LVIIESDLER AEERAELSEG KCAELEEELK TVTNNLKSLE AQAEKYSQKE DRYEEEIKVL SDKLKEAETR AE FAERSVT KLEKSIDDLE DELYAQKLKY KAISEELDHA LNDMTSI UniProtKB: Tropomyosin alpha-1 chain |
-Macromolecule #4: ADENOSINE-5'-DIPHOSPHATE
Macromolecule | Name: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 4 / Number of copies: 5 / Formula: ADP |
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Molecular weight | Theoretical: 427.201 Da |
Chemical component information | ChemComp-ADP: |
-Macromolecule #5: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 5 / Number of copies: 5 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | helical reconstruction |
Aggregation state | filament |
-Sample preparation
Concentration | 0.13 mg/mL |
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Buffer | pH: 7 |
Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 400 / Support film - Material: GOLD / Support film - topology: HOLEY |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK III |
-Electron microscopy
Microscope | TFS KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 4 / Number real images: 4171 / Average exposure time: 3.12 sec. / Average electron dose: 54.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.7000000000000001 µm / Nominal magnification: 80000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |