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- PDB-7ld0: Cryo-EM structure of ligand-free Human SARM1 -

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Basic information

Entry
Database: PDB / ID: 7ld0
TitleCryo-EM structure of ligand-free Human SARM1
ComponentsNAD(+) hydrolase SARM1
KeywordsHYDROLASE / NADase / Autoinhibition / Neurodegeneration / Allostery / NAD / Toxin / SIGNALING PROTEIN
Function / homology
Function and homology information


negative regulation of MyD88-independent toll-like receptor signaling pathway / MyD88-independent TLR4 cascade / Toll Like Receptor 3 (TLR3) Cascade / NAD catabolic process / NAD+ nucleosidase activity / ADP-ribosyl cyclase/cyclic ADP-ribose hydrolase / protein localization to mitochondrion / NAD+ nucleotidase, cyclic ADP-ribose generating / NADP+ nucleosidase activity / nervous system process ...negative regulation of MyD88-independent toll-like receptor signaling pathway / MyD88-independent TLR4 cascade / Toll Like Receptor 3 (TLR3) Cascade / NAD catabolic process / NAD+ nucleosidase activity / ADP-ribosyl cyclase/cyclic ADP-ribose hydrolase / protein localization to mitochondrion / NAD+ nucleotidase, cyclic ADP-ribose generating / NADP+ nucleosidase activity / nervous system process / Hydrolases; Glycosylases; Hydrolysing N-glycosyl compounds / regulation of dendrite morphogenesis / response to axon injury / response to glucose / signaling adaptor activity / regulation of neuron apoptotic process / TRAF6-mediated induction of TAK1 complex within TLR4 complex / Activation of IRF3, IRF7 mediated by TBK1, IKKε (IKBKE) / IKK complex recruitment mediated by RIP1 / nervous system development / microtubule / mitochondrial outer membrane / cell differentiation / axon / innate immune response / dendrite / synapse / signal transduction / mitochondrion / identical protein binding / cytosol / cytoplasm
Similarity search - Function
Sterile alpha and TIR motif-containing protein 1 / TIR domain / Toll - interleukin 1 - resistance / TIR domain profile. / Toll/interleukin-1 receptor homology (TIR) domain / Toll/interleukin-1 receptor homology (TIR) domain superfamily / SAM domain (Sterile alpha motif) / SAM domain profile. / Sterile alpha motif. / Sterile alpha motif domain ...Sterile alpha and TIR motif-containing protein 1 / TIR domain / Toll - interleukin 1 - resistance / TIR domain profile. / Toll/interleukin-1 receptor homology (TIR) domain / Toll/interleukin-1 receptor homology (TIR) domain superfamily / SAM domain (Sterile alpha motif) / SAM domain profile. / Sterile alpha motif. / Sterile alpha motif domain / Sterile alpha motif/pointed domain superfamily / Armadillo-like helical / Armadillo-type fold
Similarity search - Domain/homology
NAD(+) hydrolase SARM1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsNanson, J.D. / Gu, W. / Luo, Z. / Jia, X. / Landsberg, M.J. / Kobe, B. / Ve, T.
Funding support Australia, 6items
OrganizationGrant numberCountry
National Health and Medical Research Council (NHMRC, Australia)1107804 Australia
National Health and Medical Research Council (NHMRC, Australia)1160570 Australia
National Health and Medical Research Council (NHMRC, Australia)1071659 Australia
National Health and Medical Research Council (NHMRC, Australia)1108859 Australia
Australian Research Council (ARC)FL180100109 Australia
Australian Research Council (ARC)DE170100783 Australia
CitationJournal: Neuron / Year: 2021
Title: SARM1 is a metabolic sensor activated by an increased NMN/NAD ratio to trigger axon degeneration.
Authors: Matthew D Figley / Weixi Gu / Jeffrey D Nanson / Yun Shi / Yo Sasaki / Katie Cunnea / Alpeshkumar K Malde / Xinying Jia / Zhenyao Luo / Forhad K Saikot / Tamim Mosaiab / Veronika Masic / ...Authors: Matthew D Figley / Weixi Gu / Jeffrey D Nanson / Yun Shi / Yo Sasaki / Katie Cunnea / Alpeshkumar K Malde / Xinying Jia / Zhenyao Luo / Forhad K Saikot / Tamim Mosaiab / Veronika Masic / Stephanie Holt / Lauren Hartley-Tassell / Helen Y McGuinness / Mohammad K Manik / Todd Bosanac / Michael J Landsberg / Philip S Kerry / Mehdi Mobli / Robert O Hughes / Jeffrey Milbrandt / Bostjan Kobe / Aaron DiAntonio / Thomas Ve /
Abstract: Axon degeneration is a central pathological feature of many neurodegenerative diseases. Sterile alpha and Toll/interleukin-1 receptor motif-containing 1 (SARM1) is a nicotinamide adenine dinucleotide ...Axon degeneration is a central pathological feature of many neurodegenerative diseases. Sterile alpha and Toll/interleukin-1 receptor motif-containing 1 (SARM1) is a nicotinamide adenine dinucleotide (NAD)-cleaving enzyme whose activation triggers axon destruction. Loss of the biosynthetic enzyme NMNAT2, which converts nicotinamide mononucleotide (NMN) to NAD, activates SARM1 via an unknown mechanism. Using structural, biochemical, biophysical, and cellular assays, we demonstrate that SARM1 is activated by an increase in the ratio of NMN to NAD and show that both metabolites compete for binding to the auto-inhibitory N-terminal armadillo repeat (ARM) domain of SARM1. We report structures of the SARM1 ARM domain bound to NMN and of the homo-octameric SARM1 complex in the absence of ligands. We show that NMN influences the structure of SARM1 and demonstrate via mutagenesis that NMN binding is required for injury-induced SARM1 activation and axon destruction. Hence, SARM1 is a metabolic sensor responding to an increased NMN/NAD ratio by cleaving residual NAD, thereby inducing feedforward metabolic catastrophe and axonal demise.
History
DepositionJan 12, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 10, 2021Provider: repository / Type: Initial release
Revision 1.1Mar 17, 2021Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.name
Revision 1.2Apr 21, 2021Group: Database references / Category: citation / Item: _citation.journal_volume / _citation.page_first
Revision 1.3Mar 6, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

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Assembly

Deposited unit
A: NAD(+) hydrolase SARM1
B: NAD(+) hydrolase SARM1
C: NAD(+) hydrolase SARM1
D: NAD(+) hydrolase SARM1
E: NAD(+) hydrolase SARM1
F: NAD(+) hydrolase SARM1
G: NAD(+) hydrolase SARM1
H: NAD(+) hydrolase SARM1


Theoretical massNumber of molelcules
Total (without water)611,7728
Polymers611,7728
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: light scattering
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
NAD(+) hydrolase SARM1 / hSARM1 / NADP(+) hydrolase SARM1 / Sterile alpha and Armadillo repeat protein / Sterile alpha and ...hSARM1 / NADP(+) hydrolase SARM1 / Sterile alpha and Armadillo repeat protein / Sterile alpha and TIR motif-containing protein 1 / Sterile alpha motif domain-containing protein 2 / SAM domain-containing protein 2 / Tir-1 homolog / HsTIR


Mass: 76471.469 Da / Num. of mol.: 8
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SARM1, KIAA0524, SAMD2, SARM / Cell line (production host): HEK293T / Production host: Homo sapiens (human)
References: UniProt: Q6SZW1, ADP-ribosyl cyclase/cyclic ADP-ribose hydrolase, Hydrolases; Glycosylases; Hydrolysing N-glycosyl compounds

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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: homo-octameric SARM1 / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human) / Strain: HEK293T
Buffer solutionpH: 7.4
Buffer component
IDConc.NameBuffer-ID
125 mMHEPES1
2150 mMSodium Chloride1
30.4 %Glycerol1
40.1 mMTCEP1
SpecimenConc.: 0.27 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/2
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE

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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: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / C2 aperture diameter: 50 µm
Image recordingAverage exposure time: 4.99 sec. / Electron dose: 54 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)
Details: exposure rate of 12.88e-/pixel/second super-resolution 0.543 A/pixel 45 frames per image
EM imaging opticsEnergyfilter slit width: 20 eV

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.18.2_3874refinement
PHENIX1.18.2_3874refinement
EM software
IDNameCategory
1cryoSPARCparticle selection
4cryoSPARCCTF correction
9cryoSPARCinitial Euler assignment
10cryoSPARCfinal Euler assignment
11cryoSPARCclassification
12cryoSPARC3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C8 (8 fold cyclic)
3D reconstructionResolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 275460 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 76.97 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.005737552
ELECTRON MICROSCOPYf_angle_d0.645750728
ELECTRON MICROSCOPYf_chiral_restr0.04045848
ELECTRON MICROSCOPYf_plane_restr0.00536536
ELECTRON MICROSCOPYf_dihedral_angle_d19.66414040

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