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- PDB-7rb3: Cryo-EM structure of human binary NatC complex with a Bisubstrate... -

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Basic information

Entry
Database: PDB / ID: 7rb3
TitleCryo-EM structure of human binary NatC complex with a Bisubstrate inhibitor
Components
  • N-alpha-acetyltransferase 30
  • N-alpha-acetyltransferase 35, NatC auxiliary subunit
KeywordsTRANSFERASE / NatC / NAA30 / NAA35
Function / homology
Function and homology information


N-terminal methionine Nalpha-acetyltransferase NatC / NatC complex / smooth muscle cell proliferation / protein N-terminal-methionine acetyltransferase activity / Retrograde transport at the Trans-Golgi-Network / protein-N-terminal amino-acid acetyltransferase activity / protein stabilization / negative regulation of apoptotic process / nucleoplasm / nucleus ...N-terminal methionine Nalpha-acetyltransferase NatC / NatC complex / smooth muscle cell proliferation / protein N-terminal-methionine acetyltransferase activity / Retrograde transport at the Trans-Golgi-Network / protein-N-terminal amino-acid acetyltransferase activity / protein stabilization / negative regulation of apoptotic process / nucleoplasm / nucleus / plasma membrane / cytoplasm / cytosol
Similarity search - Function
-alpha-acetyltransferase 35, NatC auxiliary subunit / N-alpha-acetyltransferase 30-like / Mak10 subunit, NatC N(alpha)-terminal acetyltransferase / Acetyltransferase (GNAT) family / Gcn5-related N-acetyltransferase (GNAT) domain profile. / GNAT domain / Acyl-CoA N-acyltransferase
Similarity search - Domain/homology
CARBOXYMETHYL COENZYME *A / LEUCINE / METHIONINE / N-alpha-acetyltransferase 30 / N-alpha-acetyltransferase 35, NatC auxiliary subunit
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsDeng, S. / Marmorstein, R.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35 GM118090 United States
CitationJournal: Structure / Year: 2023
Title: Molecular role of NAA38 in thermostability and catalytic activity of the human NatC N-terminal acetyltransferase.
Authors: Sunbin Deng / Sarah M Gardner / Leah Gottlieb / Buyan Pan / E James Petersson / Ronen Marmorstein /
Abstract: N-terminal acetylation occurs on over 80% of human proteins and is catalyzed by a family of N-terminal acetyltransferases (NATs). All NATs contain a small catalytic subunit, while some also contain a ...N-terminal acetylation occurs on over 80% of human proteins and is catalyzed by a family of N-terminal acetyltransferases (NATs). All NATs contain a small catalytic subunit, while some also contain a large auxiliary subunit that facilitates catalysis and ribosome targeting for co-translational acetylation. NatC is one of the major NATs containing an NAA30 catalytic subunit, but uniquely contains two auxiliary subunits, large NAA35 and small NAA38. Here, we report the cryo-EM structures of human NatC (hNatC) complexes with and without NAA38, together with biochemical studies, to reveal that NAA38 increases the thermostability and broadens the substrate-specificity profile of NatC by ordering an N-terminal segment of NAA35 and reorienting an NAA30 N-terminal peptide binding loop for optimal catalysis, respectively. We also note important differences in engagement with a stabilizing inositol hexaphosphate molecule between human and yeast NatC. These studies provide new insights for the function and evolution of the NatC complex.
History
DepositionJul 5, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 11, 2023Provider: repository / Type: Initial release
Revision 1.1Jun 28, 2023Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2May 1, 2024Group: Data collection / Data processing / Category: chem_comp_atom / chem_comp_bond / em_experiment / Item: _em_experiment.entity_assembly_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
B: N-alpha-acetyltransferase 35, NatC auxiliary subunit
A: N-alpha-acetyltransferase 30
hetero molecules


Theoretical massNumber of molelcules
Total (without water)99,8975
Polymers98,7912
Non-polymers1,1063
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein N-alpha-acetyltransferase 35, NatC auxiliary subunit / Embryonic growth-associated protein homolog / Protein MAK10 homolog


Mass: 80707.625 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: NAA35, EGAP, MAK10 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q5VZE5
#2: Protein N-alpha-acetyltransferase 30 / N-acetyltransferase 12 / N-acetyltransferase MAK3 homolog / NatC catalytic subunit


Mass: 18083.293 Da / Num. of mol.: 1 / Fragment: catalytic domain
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: NAA30, C14orf35, MAK3, NAT12 / Production host: Spodoptera frugiperda (fall armyworm)
References: UniProt: Q147X3, N-terminal methionine Nalpha-acetyltransferase NatC
#3: Chemical ChemComp-CMC / CARBOXYMETHYL COENZYME *A


Mass: 825.570 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C23H38N7O18P3S / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical ChemComp-MET / METHIONINE


Type: L-peptide linking / Mass: 149.211 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C5H11NO2S
#5: Chemical ChemComp-LEU / LEUCINE


Type: L-peptide linking / Mass: 131.173 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C6H13NO2
Has ligand of interestY

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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: heterotimeric S.pombe NatC complex with a bisubstrate inhibitor and inositol hexaphosphate
Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 7
Buffer component
IDConc.NameFormulaBuffer-ID
1200 mMsodium clorideNaCl1
225 mM4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid1
31 mMDithiothreitol1
SpecimenConc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK II / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 289 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 3000 nm / Nominal defocus min: 1000 nm
Image recording
IDImaging-IDElectron dose (e/Å2)Film or detector model
111.6GATAN K3 (6k x 4k)
211.3GATAN K3 (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.20.1_4487: / Classification: refinement
EM software
IDNameVersionCategory
4CTFFINDCTF correction
7Coot0.8.9.2model fitting
9PHENIX1.17.1-3660model refinement
10cryoSPARCv2initial Euler assignment
11cryoSPARCv2final Euler assignment
12cryoSPARCv2classification
13cryoSPARCv23D reconstruction
Image processingDetails: 4222 images
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 192437 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL / Space: REAL
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0045222
ELECTRON MICROSCOPYf_angle_d0.5677144
ELECTRON MICROSCOPYf_dihedral_angle_d3.791782
ELECTRON MICROSCOPYf_chiral_restr0.04844
ELECTRON MICROSCOPYf_plane_restr0.003919

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