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- PDB-7lgq: Cyanophycin synthetase 1 from Synechocystis sp. UTEX2470 with ATP... -

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Basic information

Entry
Database: PDB / ID: 7lgq
TitleCyanophycin synthetase 1 from Synechocystis sp. UTEX2470 with ATP and 8x(Asp-Arg)-Asn
Components
  • 8x(Asp-Arg)-Asn
  • Cyanophycin synthase
KeywordsLIGASE / Cyanophycin / CphA1 / ATP-grasp / enzyme
Function / homology
Function and homology information


cyanophycin synthase (L-aspartate-adding) / cyanophycin synthase (L-arginine-adding) / cyanophycin synthetase activity (L-aspartate-adding) / cyanophycin synthetase activity (L-arginine-adding) / ATP binding / metal ion binding
Similarity search - Function
Cyanophycin synthase-like, N-terminal / Cyanophycin synthase-like N-terminal domain / Cyanophycin synthetase / ATP-grasp fold, RimK-type / RimK-like ATP-grasp domain / Mur ligase, C-terminal / Mur ligase, C-terminal domain superfamily / Mur ligase, glutamate ligase domain / Mur ligase, central / Mur-like, catalytic domain superfamily ...Cyanophycin synthase-like, N-terminal / Cyanophycin synthase-like N-terminal domain / Cyanophycin synthetase / ATP-grasp fold, RimK-type / RimK-like ATP-grasp domain / Mur ligase, C-terminal / Mur ligase, C-terminal domain superfamily / Mur ligase, glutamate ligase domain / Mur ligase, central / Mur-like, catalytic domain superfamily / Mur ligase middle domain / ATP-grasp fold / ATP-grasp fold profile.
Similarity search - Domain/homology
ADENOSINE-5'-TRIPHOSPHATE / Cyanophycin synthetase
Similarity search - Component
Biological speciesSynechocystis sp. (bacteria)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsSharon, I. / Grogg, M. / Hilvert, D. / Schmeing, T.M.
CitationJournal: Nat Chem Biol / Year: 2021
Title: Structures and function of the amino acid polymerase cyanophycin synthetase.
Authors: Itai Sharon / Asfarul S Haque / Marcel Grogg / Indrajit Lahiri / Dieter Seebach / Andres E Leschziner / Donald Hilvert / T Martin Schmeing /
Abstract: Cyanophycin is a natural biopolymer produced by a wide range of bacteria, consisting of a chain of poly-L-Asp residues with L-Arg residues attached to the β-carboxylate sidechains by isopeptide ...Cyanophycin is a natural biopolymer produced by a wide range of bacteria, consisting of a chain of poly-L-Asp residues with L-Arg residues attached to the β-carboxylate sidechains by isopeptide bonds. Cyanophycin is synthesized from ATP, aspartic acid and arginine by a homooligomeric enzyme called cyanophycin synthetase (CphA1). CphA1 has domains that are homologous to glutathione synthetases and muramyl ligases, but no other structural information has been available. Here, we present cryo-electron microscopy and X-ray crystallography structures of cyanophycin synthetases from three different bacteria, including cocomplex structures of CphA1 with ATP and cyanophycin polymer analogs at 2.6 Å resolution. These structures reveal two distinct tetrameric architectures, show the configuration of active sites and polymer-binding regions, indicate dynamic conformational changes and afford insight into catalytic mechanism. Accompanying biochemical interrogation of substrate binding sites, catalytic centers and oligomerization interfaces combine with the structures to provide a holistic understanding of cyanophycin biosynthesis.
History
DepositionJan 20, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Aug 18, 2021Provider: repository / Type: Initial release
Revision 1.1Sep 1, 2021Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Oct 6, 2021Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID
Revision 1.3Oct 23, 2024Group: Data collection / Derived calculations / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_admin / pdbx_entry_details / pdbx_modification_feature / struct_conn
Item: _em_admin.last_update / _pdbx_entry_details.has_protein_modification / _struct_conn.pdbx_leaving_atom_flag

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Structure visualization

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Assembly

Deposited unit
A: Cyanophycin synthase
B: Cyanophycin synthase
C: Cyanophycin synthase
D: Cyanophycin synthase
E: 8x(Asp-Arg)-Asn
F: 8x(Asp-Arg)-Asn
G: 8x(Asp-Arg)-Asn
H: 8x(Asp-Arg)-Asn
I: 8x(Asp-Arg)-Asn
J: 8x(Asp-Arg)-Asn
K: 8x(Asp-Arg)-Asn
L: 8x(Asp-Arg)-Asn
hetero molecules


Theoretical massNumber of molelcules
Total (without water)405,70628
Polymers401,45412
Non-polymers4,25216
Water00
1


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

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Components

#1: Protein
Cyanophycin synthase / Cyanophycin synthetase


Mass: 95758.836 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Synechocystis sp. (strain PCC 6714) (bacteria)
Strain: PCC 6714 / Gene: cphA, D082_30240 / Production host: Escherichia coli BL21(DE3) (bacteria)
References: UniProt: A0A068N621, cyanophycin synthase (L-aspartate-adding), cyanophycin synthase (L-arginine-adding)
#2: Protein/peptide
8x(Asp-Arg)-Asn


Mass: 2302.300 Da / Num. of mol.: 8 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#3: Chemical
ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE


Mass: 507.181 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Comment: ATP, energy-carrying molecule*YM
#4: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: Mg
Compound detailsEntity-2 is a Cyanophycin analog, composed of an aspartic acid backbone and arginine side groups
Has ligand of interestY
Has protein modificationY

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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: Cyanophycin synthetase 1 from Synechocystis sp. UTEX2470 with ATP and 8x(Asp-Arg)-Asn
Type: COMPLEX / Entity ID: #1-#2 / Source: MULTIPLE SOURCES
Molecular weightExperimental value: NO
Source (natural)Organism: Synechocystis sp. PCC 6714 (bacteria)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 8
SpecimenConc.: 3 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: C-flat-1.2/1.3
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: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: OTHER
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM softwareName: cryoSPARC / Category: 3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: D2 (2x2 fold dihedral)
3D reconstructionResolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 135110 / Symmetry type: POINT

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