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- EMDB-41758: Structure of C-terminal LRRK2 bound to MLi-2 (I2020T mutant) -

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Entry
Database: EMDB / ID: EMD-41758
TitleStructure of C-terminal LRRK2 bound to MLi-2 (I2020T mutant)
Map data
Sample
  • Complex: C-terminal LRRK2 (I2020T mutation) bound to MLi-2
    • Protein or peptide: Leucine-rich repeat serine/threonine-protein kinase 2
  • Ligand: (2~{R},6~{S})-2,6-dimethyl-4-[6-[5-(1-methylcyclopropyl)oxy-1~{H}-indazol-3-yl]pyrimidin-4-yl]morpholine
  • Ligand: GUANOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: water
KeywordsGtPase / kinase / inhibitors / PROTEIN BINDING
Function / homology
Function and homology information


peroxidase inhibitor activity / caveola neck / negative regulation of thioredoxin peroxidase activity by peptidyl-threonine phosphorylation / negative regulation of protein processing involved in protein targeting to mitochondrion / Wnt signalosome assembly / beta-catenin destruction complex binding / regulation of branching morphogenesis of a nerve / regulation of kidney size / regulation of neuron maturation / tangential migration from the subventricular zone to the olfactory bulb ...peroxidase inhibitor activity / caveola neck / negative regulation of thioredoxin peroxidase activity by peptidyl-threonine phosphorylation / negative regulation of protein processing involved in protein targeting to mitochondrion / Wnt signalosome assembly / beta-catenin destruction complex binding / regulation of branching morphogenesis of a nerve / regulation of kidney size / regulation of neuron maturation / tangential migration from the subventricular zone to the olfactory bulb / protein localization to endoplasmic reticulum exit site / GTP-dependent protein kinase activity / regulation of neuroblast proliferation / regulation of ER to Golgi vesicle-mediated transport / regulation of synaptic vesicle transport / negative regulation of late endosome to lysosome transport / regulation of mitochondrial depolarization / negative regulation of protein targeting to mitochondrion / positive regulation of dopamine receptor signaling pathway / regulation of lysosomal lumen pH / regulation of CAMKK-AMPK signaling cascade / amphisome / mitochondrion localization / cytoplasmic side of mitochondrial outer membrane / co-receptor binding / regulation of retrograde transport, endosome to Golgi / negative regulation of excitatory postsynaptic potential / negative regulation of autophagosome assembly / regulation of dopamine receptor signaling pathway / positive regulation of microglial cell activation / neuron projection arborization / positive regulation of synaptic vesicle endocytosis / JUN kinase kinase kinase activity / olfactory bulb development / regulation of dendritic spine morphogenesis / regulation of protein kinase A signaling / multivesicular body, internal vesicle / striatum development / protein localization to mitochondrion / cellular response to dopamine / presynaptic cytosol / positive regulation of protein autoubiquitination / endoplasmic reticulum organization / positive regulation of programmed cell death / Wnt signalosome / GTP metabolic process / regulation of canonical Wnt signaling pathway / negative regulation of protein processing / syntaxin-1 binding / regulation of reactive oxygen species metabolic process / negative regulation of GTPase activity / autolysosome / protein kinase A binding / regulation of locomotion / exploration behavior / regulation of synaptic vesicle exocytosis / PTK6 promotes HIF1A stabilization / clathrin binding / Golgi-associated vesicle / negative regulation of macroautophagy / lysosome organization / neuromuscular junction development / regulation of mitochondrial fission / intracellular distribution of mitochondria / Golgi organization / positive regulation of nitric-oxide synthase biosynthetic process / locomotory exploration behavior / microvillus / Rho protein signal transduction / cellular response to organic cyclic compound / MAP kinase kinase kinase activity / canonical Wnt signaling pathway / positive regulation of protein kinase activity / cellular response to manganese ion / endoplasmic reticulum exit site / positive regulation of autophagy / negative regulation of endoplasmic reticulum stress-induced intrinsic apoptotic signaling pathway / JNK cascade / regulation of synaptic transmission, glutamatergic / excitatory postsynaptic potential / cellular response to starvation / dendrite cytoplasm / GTPase activator activity / mitochondrion organization / regulation of membrane potential / tubulin binding / SNARE binding / neuron projection morphogenesis / negative regulation of protein phosphorylation / negative regulation of protein binding / positive regulation of protein ubiquitination / regulation of autophagy / determination of adult lifespan / calcium-mediated signaling / mitochondrial membrane / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / peptidyl-threonine phosphorylation / regulation of protein stability / positive regulation of MAP kinase activity / trans-Golgi network
Similarity search - Function
C-terminal of Roc (COR) domain / C-terminal of Roc, COR, domain / Ras of Complex, Roc, domain of DAPkinase / Roc domain profile. / Roc domain / Leucine-rich repeats, bacterial type / Leucine rich repeat / Leucine-rich repeat, typical subtype / Leucine-rich repeats, typical (most populated) subfamily / Leucine-rich repeat profile. ...C-terminal of Roc (COR) domain / C-terminal of Roc, COR, domain / Ras of Complex, Roc, domain of DAPkinase / Roc domain profile. / Roc domain / Leucine-rich repeats, bacterial type / Leucine rich repeat / Leucine-rich repeat, typical subtype / Leucine-rich repeats, typical (most populated) subfamily / Leucine-rich repeat profile. / Leucine-rich repeat / Rab subfamily of small GTPases / Leucine-rich repeat domain superfamily / Ankyrin repeat-containing domain superfamily / Armadillo-like helical / Small GTP-binding protein domain / Armadillo-type fold / WD40-repeat-containing domain superfamily / Serine/threonine-protein kinase, active site / Serine/Threonine protein kinases active-site signature. / WD40/YVTN repeat-like-containing domain superfamily / Protein kinase domain / Serine/Threonine protein kinases, catalytic domain / Protein kinase, ATP binding site / Protein kinases ATP-binding region signature. / Protein kinase domain profile. / Protein kinase domain / Protein kinase-like domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Leucine-rich repeat serine/threonine-protein kinase 2
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsSanz-Murillo M / Villagran-Suarez A / Alegrio Louro J / Leschziner A
Funding support United States, 1 items
OrganizationGrant numberCountry
Michael J. Fox FoundationASAP-000519 United States
CitationJournal: Sci Adv / Year: 2023
Title: Inhibition of Parkinson's disease-related LRRK2 by type I and type II kinase inhibitors: Activity and structures.
Authors: Marta Sanz Murillo / Amalia Villagran Suarez / Verena Dederer / Deep Chatterjee / Jaime Alegrio Louro / Stefan Knapp / Sebastian Mathea / Andres E Leschziner /
Abstract: Mutations in leucine-rich repeat kinase 2 (LRRK2) are a common cause of familial Parkinson's disease (PD) and a risk factor for the sporadic form. Increased kinase activity was shown in patients with ...Mutations in leucine-rich repeat kinase 2 (LRRK2) are a common cause of familial Parkinson's disease (PD) and a risk factor for the sporadic form. Increased kinase activity was shown in patients with both familial and sporadic PD, making LRRK2 kinase inhibitors a major focus of drug development efforts. Although much progress has been made in understanding the structural biology of LRRK2, there are no available structures of LRRK2 inhibitor complexes. To this end, we solved cryo-electron microscopy structures of LRRK2, wild-type and PD-linked mutants, bound to the LRRK2-specific type I inhibitor MLi-2 and the broad-spectrum type II inhibitor GZD-824. Our structures revealed an active-like LRRK2 kinase in the type I inhibitor complex, and an inactive DYG-out in the type II inhibitor complex. Our structural analysis also showed how inhibitor-induced conformational changes in LRRK2 are affected by its autoinhibitory N-terminal repeats. The structures provide a template for the rational development of LRRK2 kinase inhibitors covering both canonical inhibitor binding modes.
History
DepositionAug 26, 2023-
Header (metadata) releaseDec 6, 2023-
Map releaseDec 6, 2023-
UpdateDec 13, 2023-
Current statusDec 13, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_41758.png

Downloads & links

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Map

FileDownload / File: emd_41758.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.935 Å
Density
Contour LevelBy AUTHOR: 0.15
Minimum - Maximum-1.4184575 - 2.1668553
Average (Standard dev.)0.0005722314 (±0.029124314)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 374.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: #1

Fileemd_41758_additional_1.map
Projections & Slices
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Half map: #2

Fileemd_41758_half_map_1.map
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Half map: #1

Fileemd_41758_half_map_2.map
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Sample components

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Entire : C-terminal LRRK2 (I2020T mutation) bound to MLi-2

EntireName: C-terminal LRRK2 (I2020T mutation) bound to MLi-2
Components
  • Complex: C-terminal LRRK2 (I2020T mutation) bound to MLi-2
    • Protein or peptide: Leucine-rich repeat serine/threonine-protein kinase 2
  • Ligand: (2~{R},6~{S})-2,6-dimethyl-4-[6-[5-(1-methylcyclopropyl)oxy-1~{H}-indazol-3-yl]pyrimidin-4-yl]morpholine
  • Ligand: GUANOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: water

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Supramolecule #1: C-terminal LRRK2 (I2020T mutation) bound to MLi-2

SupramoleculeName: C-terminal LRRK2 (I2020T mutation) bound to MLi-2 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 137 KDa

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Macromolecule #1: Leucine-rich repeat serine/threonine-protein kinase 2

MacromoleculeName: Leucine-rich repeat serine/threonine-protein kinase 2 / type: protein_or_peptide / ID: 1
Details: Several loops are missing at the structure due to the lack of density
Number of copies: 1 / Enantiomer: LEVO / EC number: non-specific serine/threonine protein kinase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 135.589094 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: NRMKLMIVGN TGSGKTTLLQ QLMKTKKSDL GMQSATVGID VKDWPIQIRD KRKRDLVLNV WDFAGREEFY STHPHFMTQR ALYLAVYDL SKGQAEVDAM KPWLFNIKAR ASSSPVILVG THLDVSDEKQ RKACMSKITK ELLNKRGFPA IRDYHFVNAT E ESDALAKL ...String:
NRMKLMIVGN TGSGKTTLLQ QLMKTKKSDL GMQSATVGID VKDWPIQIRD KRKRDLVLNV WDFAGREEFY STHPHFMTQR ALYLAVYDL SKGQAEVDAM KPWLFNIKAR ASSSPVILVG THLDVSDEKQ RKACMSKITK ELLNKRGFPA IRDYHFVNAT E ESDALAKL RKTIINESLN FKIRDQLVVG QLIPDCYVEL EKIILSERKN VPIEFPVIDR KRLLQLVREN QLQLDENELP HA VHFLNES GVLLHFQDPA LQLSDLYFVE PKWLCKIMAQ ILTVKVEGCP KHPKGIISRR DVEKFLSKKR KFPKNYMSQY FKL LEKFQI ALPIGEEYLL VPSSLSDHRP VIELPHCENS EIIIRLYEMP YFPMGFWSRL INRLLEISPY MLSGRERALR PNRM YWRQG IYLNWSPEAY CLVGSEVLDN HPESFLKITV PSCRKGCILL GQVVDHIDSL MEEWFPGLLE IDICGEGETL LKKWA LYSF NDGEEHQKIL LDDLMKKAEE GDLLVNPDQP RLTIPISQIA PDLILADLPR NIMLNNDELE FEQAPEFLLG DGSFGS VYR AAYEGEEVAV KIFNKHTSLR LLRQELVVLC HLHHPSLISL LAAGIRPRML VMELASKGSL DRLLQQDKAS LTRTLQH RI ALHVADGLRY LHSAMIIYRD LKPHNVLLFT LYPNAAIIAK IADYGTAQYC CRMGIKTSEG TPGFRAPEVA RGNVIYNQ Q ADVYSFGLLL YDILTTGGRI VEGLKFPNEF DELEIQGKLP DPVKEYGCAP WPMVEKLIKQ CLKENPQERP TSAQVFDIL NSAELVCLTR RILLPKNVIV ECMVATHHNS RNASIWLGCG HTDRGQLSFL DLNTEGYTSE EVADSRILCL ALVHLPVEKE SWIVSGTQS GTLLVINTED GKKRHTLEKM TDSVTCLYCN SFSKQSKQKN FLLVGTADGK LAIFEDKTVK LKGAAPLKIL N IGNVSTPL MCLSESTNST ERNVMWGGCG TKIFSFSNDF TIQKLIETRT SQLFSYAAFS DSNIITVVVD TALYIAKQNS PV VEVWDKK TEKLCGLIDC VHFLREVMVK ENKESKHKMS YSGRVKTLCL QKNTALWIGT GGGHILLLDL STRRLIRVIY NFC NSVRVM MTAQLGSLKN VMLVLGYNRK NTEGTQKQKE IQSCLTVWDI NLPHEVQNLE KHIEVRKELA EKMR

UniProtKB: Leucine-rich repeat serine/threonine-protein kinase 2

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Macromolecule #2: (2~{R},6~{S})-2,6-dimethyl-4-[6-[5-(1-methylcyclopropyl)oxy-1~{H}...

MacromoleculeName: (2~{R},6~{S})-2,6-dimethyl-4-[6-[5-(1-methylcyclopropyl)oxy-1~{H}-indazol-3-yl]pyrimidin-4-yl]morpholine
type: ligand / ID: 2 / Number of copies: 1 / Formula: A1N
Molecular weightTheoretical: 379.456 Da
Chemical component information

ChemComp-A1N:
(2~{R},6~{S})-2,6-dimethyl-4-[6-[5-(1-methylcyclopropyl)oxy-1~{H}-indazol-3-yl]pyrimidin-4-yl]morpholine

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Macromolecule #3: GUANOSINE-5'-DIPHOSPHATE

MacromoleculeName: GUANOSINE-5'-DIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 1 / Formula: GDP
Molecular weightTheoretical: 443.201 Da
Chemical component information

ChemComp-GDP:
GUANOSINE-5'-DIPHOSPHATE / GDP, energy-carrying molecule*YM / Guanosine diphosphate

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Macromolecule #4: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 1 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #5: water

MacromoleculeName: water / type: ligand / ID: 5 / Number of copies: 1 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER / Water

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.822 mg/mL
BufferpH: 7.4
Component:
ConcentrationNameFormula
20.0 mMHEPES
150.0 mMSodium ChlorideNaClSodium chloride
2.5 mMMagnesium ChlorideMgCl2
5.0 %GlycerolGlyOH
20.0 uMGDP
0.5 mMTCEP
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV
Details: Blot force = 3 Blot time = 4 seconds Wait time = 20 seconds.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 4.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 130000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Number grids imaged: 1 / Number real images: 10050 / Average exposure time: 11.0 sec. / Average electron dose: 55.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 628666
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

6vp7

Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.0)
Final reconstructionApplied symmetry - Point group: D2 (2x2 fold dihedral) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.0) / Number images used: 380487
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6vp7


Chain - Chain ID: A / Chain - Residue range: 1333-2527 / Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-8tzg:
Structure of C-terminal LRRK2 bound to MLi-2 (I2020T mutant)

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