[English] 日本語
Yorodumi
- PDB-3dtp: Tarantula heavy meromyosin obtained by flexible docking to Tarant... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 3dtp
TitleTarantula heavy meromyosin obtained by flexible docking to Tarantula muscle thick filament Cryo-EM 3D-MAP
Components
  • (Myosin 2 heavy chain chimera of smooth and cardiac muscle) x 2
  • Myosin regulatory light chain
  • Smooth muscle myosin essential light chain
KeywordsCONTRACTILE PROTEIN / MUSCLE PROTEIN / SMOOTH MUSCLE / MYOSIN SUBFRAGMENT 2 / HEAVY MEROMYOSIN / ESSENTIAL LIGHT CHAIN / REGULATORY LIGHT CHAIN / MOTOR PROTEIN / COILED-COIL
Function / homology
Function and homology information


myosin II filament / regulation of slow-twitch skeletal muscle fiber contraction / regulation of the force of skeletal muscle contraction / myofibril assembly / myosin light chain binding / muscle myosin complex / elastic fiber assembly / adult heart development / regulation of the force of heart contraction / myosin II complex ...myosin II filament / regulation of slow-twitch skeletal muscle fiber contraction / regulation of the force of skeletal muscle contraction / myofibril assembly / myosin light chain binding / muscle myosin complex / elastic fiber assembly / adult heart development / regulation of the force of heart contraction / myosin II complex / skeletal muscle myosin thick filament assembly / transition between fast and slow fiber / actomyosin structure organization / cardiac muscle hypertrophy in response to stress / myosin II binding / actin-dependent ATPase activity / cardiac muscle fiber development / myosin filament / skeletal muscle contraction / striated muscle contraction / microfilament motor activity / actomyosin / muscle filament sliding / myosin complex / structural constituent of muscle / smooth muscle contraction / myofibril / myosin heavy chain binding / ventricular cardiac muscle tissue morphogenesis / ATP metabolic process / cardiac muscle contraction / stress fiber / muscle contraction / sarcomere / ADP binding / regulation of heart rate / Z disc / actin filament binding / actin binding / calmodulin binding / ATPase activity / calcium ion binding / protein heterodimerization activity / magnesium ion binding / ATP binding / cytosol
EF-hand domain pair / Myosin S1 fragment, N-terminal / IQ motif, EF-hand binding site / Myosin head, motor domain / EF-hand domain / Myosin tail / Myosin, N-terminal, SH3-like / EF-hand domain pair / EF-Hand 1, calcium-binding site / Myosin IQ motif-containing domain superfamily ...EF-hand domain pair / Myosin S1 fragment, N-terminal / IQ motif, EF-hand binding site / Myosin head, motor domain / EF-hand domain / Myosin tail / Myosin, N-terminal, SH3-like / EF-hand domain pair / EF-Hand 1, calcium-binding site / Myosin IQ motif-containing domain superfamily / P-loop containing nucleoside triphosphate hydrolase / Kinesin motor domain superfamily / Myosin head (motor domain) / Myosin N-terminal SH3-like domain / EF-hand domain
Myosin light polypeptide 6 / Myosin-11 / Myosin-7 / Myosin II regulatory light chain
Biological speciesGallus gallus (chicken)
Homo sapiens (human)
Avicularia avicularia (pinktoe tarantula)
MethodELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 20 Å
AuthorsAlamo, L. / Wriggers, W. / Pinto, A. / Bartoli, F. / Salazar, L. / Zhao, F.Q. / Craig, R. / Padron, R.
Citation
Journal: J. Mol. Biol. / Year: 2008
Title: Three-dimensional reconstruction of tarantula myosin filaments suggests how phosphorylation may regulate myosin activity.
Authors: Lorenzo Alamo / Willy Wriggers / Antonio Pinto / Fulvia Bártoli / Leiria Salazar / Fa-Qing Zhao / Roger Craig / Raúl Padrón /
Abstract: Muscle contraction involves the interaction of the myosin heads of the thick filaments with actin subunits of the thin filaments. Relaxation occurs when this interaction is blocked by molecular ...Muscle contraction involves the interaction of the myosin heads of the thick filaments with actin subunits of the thin filaments. Relaxation occurs when this interaction is blocked by molecular switches on these filaments. In many muscles, myosin-linked regulation involves phosphorylation of the myosin regulatory light chains (RLCs). Electron microscopy of vertebrate smooth muscle myosin molecules (regulated by phosphorylation) has provided insight into the relaxed structure, revealing that myosin is switched off by intramolecular interactions between its two heads, the free head and the blocked head. Three-dimensional reconstruction of frozen-hydrated specimens revealed that this asymmetric head interaction is also present in native thick filaments of tarantula striated muscle. Our goal in this study was to elucidate the structural features of the tarantula filament involved in phosphorylation-based regulation. A new reconstruction revealed intra- and intermolecular myosin interactions in addition to those seen previously. To help interpret the interactions, we sequenced the tarantula RLC and fitted an atomic model of the myosin head that included the predicted RLC atomic structure and an S2 (subfragment 2) crystal structure to the reconstruction. The fitting suggests one intramolecular interaction, between the cardiomyopathy loop of the free head and its own S2, and two intermolecular interactions, between the cardiac loop of the free head and the essential light chain of the blocked head and between the Leu305-Gln327 interaction loop of the free head and the N-terminal fragment of the RLC of the blocked head. These interactions, added to those previously described, would help switch off the thick filament. Molecular dynamics simulations suggest how phosphorylation could increase the helical content of the RLC N-terminus, weakening these interactions, thus releasing both heads and activating the thick filament.
#1: Journal: J. Mol. Biol. / Year: 2003
Title: Refined model of the 10S conformation of smooth muscle myosin by cryo-electron microscopy 3D image reconstruction.
Authors: Jun Liu / Thomas Wendt / Dianne Taylor / Kenneth Taylor /
Abstract: The actin-activated ATPase activity of smooth muscle myosin and heavy meromyosin (smHMM) is regulated by phosphorylation of the regulatory light chain (RLC). Complete regulation requires two intact ...The actin-activated ATPase activity of smooth muscle myosin and heavy meromyosin (smHMM) is regulated by phosphorylation of the regulatory light chain (RLC). Complete regulation requires two intact myosin heads because single-headed myosin subfragments are always active. 2D crystalline arrays of the 10S form of intact myosin, which has a dephosphorylated RLC, were produced on a positively charged lipid monolayer and imaged in 3D at 2.0 nm resolution by cryo-electron microscopy of frozen, hydrated specimens. An atomic model of smooth muscle myosin was constructed from the X-ray structures of the smooth muscle myosin motor domain and essential light chain and a homology model of the RLC was produced based on the skeletal muscle S1 structure. The initial model of the 10S myosin, based on the previous reconstruction of smHMM, was subjected to real space refinement to obtain a quantitative fit to the density. The smHMM was likewise refined and both refined models reveal the same asymmetric interaction between the upper 50 kDa domain of the "blocked" head and parts of the catalytic, converter domains and the essential light chain of the "free" head observed previously. This observation suggests that this interaction is not simply due to crystallographic packing but is enforced by elements of the myosin heads. The 10S reconstruction shows additional alpha-helical coiled-coil not seen in the earlier smHMM reconstruction, but the location of one segment of S2 is the same in both.
#2: Journal: Proc. Natl. Acad. Sci. U.S.A. / Year: 2006
Title: Crystal structures of human cardiac beta-myosin II S2-Delta provide insight into the functional role of the S2 subfragment.
Authors: Wulf Blankenfeldt / Nicolas H Thomä / John S Wray / Mathias Gautel / Ilme Schlichting /
Abstract: Myosin II is the major component of the muscle thick filament. It consists of two N-terminal S1 subfragments ("heads") connected to a long dimeric coiled-coil rod. The rod is in itself twofold ...Myosin II is the major component of the muscle thick filament. It consists of two N-terminal S1 subfragments ("heads") connected to a long dimeric coiled-coil rod. The rod is in itself twofold symmetric, but in the filament, the two heads point away from the filament surface and are therefore not equivalent. This breaking of symmetry requires the initial section of the rod, subfragment 2 (S2), to be relatively flexible. S2 is an important functional element, involved in various mechanisms by which the activity of smooth and striated muscle is regulated. We have determined crystal structures of the 126 N-terminal residues of S2 from human cardiac beta-myosin II (S2-Delta), of both WT and the disease-associated E924K mutant. S2-Delta is a straight parallel dimeric coiled coil, but the N terminus of one chain is disordered in WT-S2-Delta due to crystal contacts, indicative of unstable local structure. Bulky noncanonical side chains pack into a/d positions of S2-Delta's N terminus, leading to defined local asymmetry and axial stagger, which could induce nonequivalence of the S1 subfragments. Additionally, S2 possesses a conserved charge distribution with three prominent rings of negative potential within S2-Delta, the first of which may provide a binding interface for the "blocked head" of smooth muscle myosin in the OFF state. The observation that many disease-associated mutations affect the second negatively charged ring further suggests that charge interactions play an important role in regulation of cardiac muscle activity through myosin-binding protein C.
#3: Journal: Cell / Year: 1999
Title: Atomic structure of scallop myosin subfragment S1 complexed with MgADP: a novel conformation of the myosin head.
Authors: A Houdusse / V N Kalabokis / D Himmel / A G Szent-Györgyi / C Cohen /
Abstract: The crystal structure of a proteolytic subfragment from scallop striated muscle myosin, complexed with MgADP, has been solved at 2.5 A resolution and reveals an unusual conformation of the myosin ...The crystal structure of a proteolytic subfragment from scallop striated muscle myosin, complexed with MgADP, has been solved at 2.5 A resolution and reveals an unusual conformation of the myosin head. The converter and the lever arm are in very different positions from those in either the pre-power stroke or near-rigor state structures; moreover, in contrast to these structures, the SH1 helix is seen to be unwound. Here we compare the overall organization of the myosin head in these three states and show how the conformation of three flexible "joints" produces rearrangements of the four major subdomains in the myosin head with different bound nucleotides. We believe that this novel structure represents one of the prehydrolysis ("ATP") states of the contractile cycle in which the myosin heads stay detached from actin.
Validation Report
SummaryFull reportAbout validation report
History
DepositionJul 15, 2008Deposition site: RCSB / Processing site: PDBJ
Revision 1.0Oct 7, 2008Provider: repository / Type: Initial release
Revision 1.1Jul 13, 2011Group: Version format compliance
Revision 1.2Jul 22, 2015Group: Source and taxonomy
Revision 1.3Dec 14, 2016Group: Other

-
Structure visualization

Movie
  • Biological unit as representative helical assembly
  • Imaged by Jmol
  • Download
  • Deposited structure unit
  • Imaged by Jmol
  • Download
  • Superimposition on EM map
  • EMDB-1535
  • Imaged by UCSF Chimera
  • Download
  • Superimposition on EM map
  • EMDB-1535
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
A: Myosin 2 heavy chain chimera of smooth and cardiac muscle
B: Myosin 2 heavy chain chimera of smooth and cardiac muscle
C: Smooth muscle myosin essential light chain
D: Smooth muscle myosin essential light chain
E: Myosin regulatory light chain
F: Myosin regulatory light chain


Theoretical massNumber of molelcules
Total (without water)301,0676
Polymers301,0676
Non-polymers00
Water0
1
A: Myosin 2 heavy chain chimera of smooth and cardiac muscle
B: Myosin 2 heavy chain chimera of smooth and cardiac muscle
C: Smooth muscle myosin essential light chain
D: Smooth muscle myosin essential light chain
E: Myosin regulatory light chain
F: Myosin regulatory light chain
x 36


Theoretical massNumber of molelcules
Total (without water)10,838,410216
Polymers10,838,410216
Non-polymers00
Water0
TypeNameSymmetry operationNumber
helical symmetry operation36
2


  • Idetical with deposited unit in distinct coordinate
  • helical asymmetric unit
TypeNameSymmetry operationNumber
helical symmetry operation1
3


  • Idetical with deposited unit in distinct coordinate
  • helical asymmetric unit, std helical frame
TypeNameSymmetry operationNumber
transform to helical frame1
SymmetryHelical symmetry: (Circular symmetry: 4 / Dyad axis: no / N subunits divisor: 1 / Num. of operations: 36 / Rise per n subunits: 145 Å / Rotation per n subunits: 30 °)

-
Components

#1: Protein/peptide Myosin 2 heavy chain chimera of smooth and cardiac muscle / Myosin heavy chain / gizzard smooth muscle / cardiac muscle beta isoform / MyHC-beta / Myosin heavy chain slow isoform / MyHC-slow


Mass: 111851.070 Da / Num. of mol.: 1 / Fragment: SUBFRAGMENT 1(S1), DELTA-S2 (residues 2-972)
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Gallus gallus, Homo sapiens / Plasmid: PVL1392, PET / Cell line (production host): SF9
Production host: Spodoptera frugiperda, Escherichia coli BL21
Strain (production host): BL21 / References: UniProt: P10587, UniProt: P12883
#2: Protein/peptide Myosin 2 heavy chain chimera of smooth and cardiac muscle / Myosin heavy chain / gizzard smooth muscle / cardiac muscle beta isoform / MyHC-beta / Myosin heavy chain slow isoform / MyHC-slow


Mass: 112051.328 Da / Num. of mol.: 1 / Fragment: SUBFRAGMENT 1(S1), DELTA-S2 (residues 2-974)
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Gallus gallus, Homo sapiens / Plasmid: PVL1392, PET / Cell line (production host): SF9
Production host: Spodoptera frugiperda, Escherichia coli BL21
Strain (production host): BL21 / References: UniProt: P10587, UniProt: P12883
#3: Protein/peptide Smooth muscle myosin essential light chain / ELC / Myosin light chain alkali smooth-muscle/non-muscle isoforms / G2 catalytic / LC17-GI / LC17-NM


Mass: 16873.025 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Gallus gallus (chicken) / Plasmid: PVL1392 / Cell line (production host): SF9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P02607
#4: Protein/peptide Myosin regulatory light chain / RLC


Mass: 21709.250 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Avicularia avicularia (pinktoe tarantula) / Tissue: leg muscle / References: UniProt: B4XT43*PLUS
Sequence detailsTHE CONFLICT LISTED FOR CHAINS A AND B, RESIDUE 2 ARE CONSISTENT WITH PDB ENTRY 1BR1 AND 1I84. THE ...THE CONFLICT LISTED FOR CHAINS A AND B, RESIDUE 2 ARE CONSISTENT WITH PDB ENTRY 1BR1 AND 1I84. THE SEQUENCE OF THE HEAVY CHAIN (CHICKEN) STRUCTURE REPORTED HERE DIFFERS FROM THAT REPORTED IN THE SWISS-PROT DATABASE (ID: P10587), THAT RESIDUE SER 2 WAS CHANGED BY ALA AND DECLARED AS CLONING ARTIFACT IN PDB 1BR1. PEPTIDE CHAIN DESIGNATIONS: THE TERMS "BLOCKED" AND "FREE" REFER TO THE CONFORMATIONS OF THE TWO S1 MYOSIN HEADS. "FREE" MYOSIN HEAD MYOSIN HEAVY CHAIN S1 PLUS S2 FRAGMENT IS CHAIN A ELC IS CHAIN C RLC IS CHAIN E "BLOCKED" MYOSIN HEAD MYOSIN HEAVY CHAIN S1 PLUS S2 FRAGMENT IS CHAIN B ELC IS CHAIN D RLC IS CHAIN F SEQUENCE GAPS IN THE MOLECULAR MODEL: HEAVY CHAIN UNP-P10587 CHAIN A,B: 1, 205-210, 452-457, 635-655, 853-1185 HEAVY CHAIN S2 FRAGMENT UNP-P12883 CHAIN A: 1-841, 962-1935 HEAVY CHAIN S2 FRAGMENT UNP-P12883 CHAIN B: 1-841, 964-1935 ELC UNP-P02607 CHAIN C,D: 1-2

-
Experimental details

-
Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: FILAMENT / 3D reconstruction method: helical reconstruction

-
Sample preparation

ComponentName: Myosin thick filaments from Tarantula striated muscle / Type: COMPLEX
Details: Polymer of a multiple myosin assembled over a paramyosin core. MODEL BUILDING: the atomic model consists of two s1 heads and a segment of s2. each heavy meromyosin consists of a chimera built by chicken smooth muscle heavy chain (1i84) for s1 plus human cardiac muscle (2fxm) for s2 (chains a,b) and two light chains, the chicken smooth muscle (1i84) for elc (chains c,d) and a homology model based on 1br1 of the tarantula skeletal muscle rlc sequence (chains e,f), this model was flexible fitted to a tarantula 3d map (emd-1535)
Buffer solutionName: 100mM NaCl, 3mM MgCl2, 1mM EGTA, 5mM PIPES, 5mM NaH2PO4, 1mM NaN3
pH: 7
Details: 100mM NaCl, 3mM MgCl2, 1mM EGTA, 5mM PIPES, 5mM NaH2PO4, 1mM NaN3
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: Holey carbon grids, 400 mesh
VitrificationDetails: Plunging in a liquid ethane. Blotting was performed from one side of the grid till a thin sample film on it using Whatman No 42 filter paper, then the grid was immediately plunged under gravity into liquid ethane cooled by liquid nitrogen. Grids were stored under liquid nitrogen.

-
Electron microscopy imaging

MicroscopyModel: FEI/PHILIPS CM120T / Date: Sep 19, 2001
Details: Holey carbon grids Cryo preserved in Liquid ethane were observed in a Philips CM120 electron microscope under low dose conditions. Only filaments on thin carbon over holes were photographed
Electron gunElectron source: LAB6 / Accelerating voltage: 120 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 35000 X / Calibrated magnification: 35000 X / Nominal defocus max: 1950 nm / Nominal defocus min: 1950 nm / Cs: 2 mm
Specimen holderTemperature: 88 K
Image scansNum. digital images: 465

-
Processing

EM software

Category: model fitting

IDNameVersion
1Situs3.2
2X-PLOR3.851
CTF correctionDetails: no corrected
3D reconstructionMethod: Single particle reconstruction with a modification of the IHRSR method
Resolution: 20 Å / Num. of particles: 15504 / Nominal pixel size: 2.48 Å / Actual pixel size: 2.48 Å / Magnification calibration: Tropomyosin paracrystal
Details: three-dimensional single particle reconstruction was carried out by a modification of the ihrsr method, using spider. low-dose electron micrographs of 1008 frozen-hydrated thick filaments halves were digitized at 0.248 nm per pixel using a nikon super coolscan 8000 ed scanner. filaments were aligned with the bare zone at the top, to ensure correct polarity in subsequent steps. a total of 15,504 segments, each 62 nm long, with an overlap of 55.8 nm, and containing aprox. 40,000 unique pairs of interacting myosin heads went into the reconstruction. as an initial reference model we used the tarantula negatively stained 3d-map, which was axially rotated, axially shifted and also out of plane tilted up to plus-minus12deg. for projection matching, giving a total of 4,095 projections (13 tilted projections plus-minus 12 deg. every 2 deg., 45 reference rotated projections (0-90 deg., every 2 deg. rotation angle), and 7 image axial shifts of 2.2 nm. the resulting 3d-map combines about 10,700 out of 15,504 filament segments, a yield of 69 percent of included segments.
Symmetry type: HELICAL
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL / Target criteria: Constrained Molecular Dynamics
Details: METHOD--Flexible Fitting REFINEMENT PROTOCOL--Custom skeleton of 31 positional markers
Atomic model building
IDPDB-ID3D fitting-ID
11I84

1i84

1
22FXM

2fxm

1
31B7T

1b7t

1
Refinement stepCycle: LAST
ProteinNucleic acidLigandSolventTotal
Num. atoms20580 0 0 0 20580

+
About Yorodumi

-
News

-
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force. (see PDBe EMDB page)
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.: Q: What is "EMD"? / ID/Accession-code notation in Yorodumi/EM Navigator

External links: EMDB at PDBe / Contact to PDBj

-
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary. This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated. See below links for details.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software). Now, EM Navigator and Yorodumi are based on the updated data.

External links: wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

+
Jun 16, 2017. Omokage search with filter

Omokage search with filter

  • Result of Omokage search can be filtered by keywords and the database types

Related info.: Omokage search

+
Sep 15, 2016. EM Navigator & Yorodumi renewed

EM Navigator & Yorodumi renewed

  • New versions of EM Navigator and Yorodumi started

Related info.: Changes in new EM Navigator and Yorodumi

+
Aug 31, 2016. New EM Navigator & Yorodumi

New EM Navigator & Yorodumi

  • In 15th Sep 2016, the development versions of EM Navigator and Yorodumi will replace the official versions.
  • Current version will continue as 'legacy version' for some time.

Related info.: Changes in new EM Navigator and Yorodumi / EM Navigator / Yorodumi

Read more

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.

Related info.: EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more