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- PDB-9ewy: CryoEM structure of human MICAL1 -

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Basic information

Entry
Database: PDB / ID: 9ewy
TitleCryoEM structure of human MICAL1
Components[F-actin]-monooxygenase MICAL1
KeywordsSIGNALING PROTEIN / MICAL / actin / actin depolymerization / axon guidance plexin / Rab
Function / homology
Function and homology information


hippocampal mossy fiber expansion / : / F-actin monooxygenase / F-actin monooxygenase activity / NAD(P)H oxidase (H2O2-forming) / sulfur oxidation / regulation of regulated secretory pathway / NAD(P)H oxidase H2O2-forming activity / oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen, NAD(P)H as one donor, and incorporation of one atom of oxygen / actin filament depolymerization ...hippocampal mossy fiber expansion / : / F-actin monooxygenase / F-actin monooxygenase activity / NAD(P)H oxidase (H2O2-forming) / sulfur oxidation / regulation of regulated secretory pathway / NAD(P)H oxidase H2O2-forming activity / oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen, NAD(P)H as one donor, and incorporation of one atom of oxygen / actin filament depolymerization / intermediate filament / actin filament bundle assembly / intercellular bridge / cytoskeleton organization / FAD binding / actin filament / monooxygenase activity / small GTPase binding / SH3 domain binding / actin filament binding / actin cytoskeleton / actin binding / Factors involved in megakaryocyte development and platelet production / midbody / endosome membrane / ciliary basal body / cilium / negative regulation of apoptotic process / protein kinase binding / signal transduction / nucleoplasm / metal ion binding / plasma membrane / cytosol / cytoplasm
Similarity search - Function
bMERB domain / Bivalent Mical/EHBP Rab binding domain / bMERB domain profile. / DUF3585 / : / LIM zinc-binding domain signature. / LIM domain / Zinc-binding domain present in Lin-11, Isl-1, Mec-3. / Zinc finger, LIM-type / LIM domain profile. ...bMERB domain / Bivalent Mical/EHBP Rab binding domain / bMERB domain profile. / DUF3585 / : / LIM zinc-binding domain signature. / LIM domain / Zinc-binding domain present in Lin-11, Isl-1, Mec-3. / Zinc finger, LIM-type / LIM domain profile. / FAD-binding domain / FAD binding domain / Calponin homology domain / Calponin homology (CH) domain / Calponin homology domain / CH domain superfamily / Calponin homology (CH) domain profile. / FAD/NAD(P)-binding domain superfamily
Similarity search - Domain/homology
FLAVIN-ADENINE DINUCLEOTIDE / [F-actin]-monooxygenase MICAL1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsSchrofel, A. / Pinkas, D. / Novacek, J. / Rozbesky, D.
Funding support Czech Republic, 1items
OrganizationGrant numberCountry
Czech Science Foundation21-27204M Czech Republic
CitationJournal: Nat Commun / Year: 2024
Title: Structural basis of MICAL autoinhibition.
Authors: Matej Horvath / Adam Schrofel / Karolina Kowalska / Jan Sabo / Jonas Vlasak / Farahdokht Nourisanami / Margarita Sobol / Daniel Pinkas / Krystof Knapp / Nicola Koupilova / Jiri Novacek / ...Authors: Matej Horvath / Adam Schrofel / Karolina Kowalska / Jan Sabo / Jonas Vlasak / Farahdokht Nourisanami / Margarita Sobol / Daniel Pinkas / Krystof Knapp / Nicola Koupilova / Jiri Novacek / Vaclav Veverka / Zdenek Lansky / Daniel Rozbesky
Abstract: MICAL proteins play a crucial role in cellular dynamics by binding and disassembling actin filaments, impacting processes like axon guidance, cytokinesis, and cell morphology. Their cellular activity ...MICAL proteins play a crucial role in cellular dynamics by binding and disassembling actin filaments, impacting processes like axon guidance, cytokinesis, and cell morphology. Their cellular activity is tightly controlled, as dysregulation can lead to detrimental effects on cellular morphology. Although previous studies have suggested that MICALs are autoinhibited, and require Rab proteins to become active, the detailed molecular mechanisms remained unclear. Here, we report the cryo-EM structure of human MICAL1 at a nominal resolution of 3.1 Å. Structural analyses, alongside biochemical and functional studies, show that MICAL1 autoinhibition is mediated by an intramolecular interaction between its N-terminal catalytic and C-terminal coiled-coil domains, blocking F-actin interaction. Moreover, we demonstrate that allosteric changes in the coiled-coil domain and the binding of the tripartite assembly of CH-L2α1-LIM domains to the coiled-coil domain are crucial for MICAL activation and autoinhibition. These mechanisms appear to be evolutionarily conserved, suggesting a potential universality across the MICAL family.
History
DepositionApr 5, 2024Deposition site: PDBE / Processing site: PDBE
Revision 1.0Nov 20, 2024Provider: repository / Type: Initial release

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Structure visualization

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MolmilJmol/JSmol

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Assembly

Deposited unit
A: [F-actin]-monooxygenase MICAL1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)118,9314
Polymers118,0151
Non-polymers9163
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein [F-actin]-monooxygenase MICAL1 / Molecule interacting with CasL protein 1 / MICAL-1 / NEDD9-interacting protein with calponin ...Molecule interacting with CasL protein 1 / MICAL-1 / NEDD9-interacting protein with calponin homology and LIM domains


Mass: 118014.734 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: MICAL1, MICAL, NICAL / Production host: Spodoptera frugiperda (fall armyworm)
References: UniProt: Q8TDZ2, F-actin monooxygenase, NAD(P)H oxidase (H2O2-forming)
#2: Chemical ChemComp-FAD / FLAVIN-ADENINE DINUCLEOTIDE


Mass: 785.550 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C27H33N9O15P2 / Comment: FAD*YM
#3: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Zn
Has ligand of interestN
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: MICAL1 with the FAD cofactor / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
10.015 MTris1
20.15 Msodium chlorideNaCl1
30.002 MDTT1
40.003 MCHAPSO1
SpecimenConc.: 3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 90 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELD / Nominal magnification: 105000 X / Calibrated magnification: 59981 X / Nominal defocus max: 2800 nm / Nominal defocus min: 1400 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN
Image recordingAverage exposure time: 2.7 sec. / Electron dose: 21 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of real images: 59961
EM imaging opticsEnergyfilter slit width: 20 eV

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Processing

EM software
IDNameVersionCategory
1cryoSPARC4.4.1particle selection
2SerialEMimage acquisition
4cryoSPARC4.4.1CTF correction
7UCSF ChimeraX1.3model fitting
11cryoSPARC4.4.1classification
12RELION43D reconstruction
19PHENIX1.21-5207model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 6256757
3D reconstructionResolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 84863 / Symmetry type: POINT
Atomic model buildingB value: 77.02 / Protocol: RIGID BODY FIT / Space: REAL
Atomic model building

3D fitting-ID: 1 / Source name: PDB / Type: experimental model

IDPDB-IDPdb chain-IDAccession codeChain-IDInitial refinement model-ID
12BRY

2bry

A2BRYA1
24TXI

4txi

4TXI2
35LE0

5le0

5LE03
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 76.71 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0036634
ELECTRON MICROSCOPYf_angle_d0.48418989
ELECTRON MICROSCOPYf_chiral_restr0.0382989
ELECTRON MICROSCOPYf_plane_restr0.00361157
ELECTRON MICROSCOPYf_dihedral_angle_d5.4847890

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