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- EMDB-37339: Cryo-EM structure of ClassIII Lanthipeptide modification enzyme P... -

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Basic information

Entry
Database: EMDB / ID: EMD-37339
TitleCryo-EM structure of ClassIII Lanthipeptide modification enzyme PneKC with chain A bounded to substrate PneA and GTP.
Map data
Sample
  • Complex: Dimeric structure of Class III lanthipeptide modification enzyme PneKC with one protomer in complex with substrate PneA and GTP
    • Protein or peptide: Protein kinase domain-containing protein
    • Protein or peptide: PneA LP
  • Ligand: GUANOSINE-5'-TRIPHOSPHATE
  • Ligand: PHOSPHATE ION
KeywordsLanthibiotic / RiPPs / LanKC / CryoEM / Antimicrobial peptides / Antiviral peptides. / ANTIMICROBIAL PROTEIN
Function / homology
Function and homology information


peptide modification / protein kinase activity / ATP binding
Similarity search - Function
Lanthionine synthetase C-like protein / Lanthionine synthetase C-like / : / Protein kinase domain profile. / Protein kinase domain / Protein kinase-like domain superfamily
Similarity search - Domain/homology
Protein kinase domain-containing protein
Similarity search - Component
Biological speciesStreptococcus pneumoniae (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.98 Å
AuthorsLi Y / Luo M / Shao K / Li J / Li Z
Funding support Singapore, 1 items
OrganizationGrant numberCountry
Ministry of Education (MoE, Singapore) Singapore
CitationJournal: Nat Commun / Year: 2024
Title: Mechanistic insights into lanthipeptide modification by a distinct subclass of LanKC enzyme that forms dimers.
Authors: Yifan Li / Kai Shao / Zhaoxing Li / Kongfu Zhu / Bee Koon Gan / Jian Shi / Yibei Xiao / Min Luo /
Abstract: Naturally occurring lanthipeptides, peptides post-translationally modified by various enzymes, hold significant promise as antibiotics. Despite extensive biochemical and structural studies, the ...Naturally occurring lanthipeptides, peptides post-translationally modified by various enzymes, hold significant promise as antibiotics. Despite extensive biochemical and structural studies, the events preceding peptide modification remain poorly understood. Here, we identify a distinct subclass of lanthionine synthetase KC (LanKC) enzymes with distinct structural and functional characteristics. We show that PneKC, a member of this subclass, forms a dimer and possesses GTPase activity. Through three cryo-EM structures of PneKC, we illustrate different stages of peptide PneA binding, from initial recognition to full binding. Our structures show the kinase domain complexed with the PneA core peptide and GTPγS, a phosphate-bound lyase domain, and an unconventional cyclase domain. The leader peptide of PneA interact with a gate loop, transitioning from an extended to a helical conformation. We identify a dimerization hot spot and propose a "negative cooperativity" mechanism toggling the enzyme between tense and relaxed conformation. Additionally, we identify an important salt bridge in the cyclase domain, differing from those in in conventional cyclase domains. These residues are highly conserved in the LanKC subclass and are part of two signature motifs. These results unveil potential differences in lanthipeptide modification enzymes assembly and deepen our understanding of allostery in these multifunctional enzymes.
History
DepositionAug 30, 2023-
Header (metadata) releaseAug 28, 2024-
Map releaseAug 28, 2024-
UpdateAug 28, 2024-
Current statusAug 28, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_37339.png

Downloads & links

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Map

FileDownload / File: emd_37339.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.86 Å/pix.
x 256 pix.
= 219.648 Å		0.86 Å/pix.
x 256 pix.
= 219.648 Å		0.86 Å/pix.
x 256 pix.
= 219.648 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.858 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.212
Minimum - Maximum-0.9436445 - 1.3601797
Average (Standard dev.)0.002924332 (±0.047391716)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 219.648 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_37339_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_37339_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Dimeric structure of Class III lanthipeptide modification enzyme ...

EntireName: Dimeric structure of Class III lanthipeptide modification enzyme PneKC with one protomer in complex with substrate PneA and GTP
Components
  • Complex: Dimeric structure of Class III lanthipeptide modification enzyme PneKC with one protomer in complex with substrate PneA and GTP
    • Protein or peptide: Protein kinase domain-containing protein
    • Protein or peptide: PneA LP
  • Ligand: GUANOSINE-5'-TRIPHOSPHATE
  • Ligand: PHOSPHATE ION

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Supramolecule #1: Dimeric structure of Class III lanthipeptide modification enzyme ...

SupramoleculeName: Dimeric structure of Class III lanthipeptide modification enzyme PneKC with one protomer in complex with substrate PneA and GTP
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Streptococcus pneumoniae (bacteria)
Molecular weightTheoretical: 210 KDa

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Macromolecule #1: Protein kinase domain-containing protein

MacromoleculeName: Protein kinase domain-containing protein / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Streptococcus pneumoniae (bacteria)
Molecular weightTheoretical: 100.334922 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: YNFNLEHPFF FTNNDYSTDT SIKYQASLPF NWHEVMNNDE WVYQYPIGKF VERQGWKIHI SSEYNSSHEL LQDVAKICHE MRIPFKHLS TEDKFIMRNG KLVSRGFSGK FITCYPNQNE LESVLQRLES ALKQYNGPYI LSDKRWDEAP IYLRYGVFRP S RDDEKKVA ...String:
YNFNLEHPFF FTNNDYSTDT SIKYQASLPF NWHEVMNNDE WVYQYPIGKF VERQGWKIHI SSEYNSSHEL LQDVAKICHE MRIPFKHLS TEDKFIMRNG KLVSRGFSGK FITCYPNQNE LESVLQRLES ALKQYNGPYI LSDKRWDEAP IYLRYGVFRP S RDDEKKVA IDELIVGDEV VKDERLPVFK IPKGIVPPDF LNKWLDKKDK KQGDFPFIID NAIRFSNSGG IYNARLKEDG KK IILKEAR PYTGLGFDGT YSSEKLASEC KALKILNEWS EAPKIYWHGK IWEHTFLGIE HMKGVPLNRW VTNNFPLYEV VDK TKDYLL RVSKIVEKLI DLTNKFHSEN VYHQDLHLGN ILVKDEDEIS IIDWEQAVFS NDEKVVHKVA APGFRAWRET LPSE IDWYG IRQIAHYLYM PLVTTSDLTY NYVSQTRIEG KKLFESLGYT REHIDYVESL LSYLDSKCPQ IENISRKKVL KPMHE IRTI ESEQDIQDFI IKLLRGFTLT YGQWRKEFQS RFFPVHYYGL NFNQGIAFSD LAILWSYQQL AKKVKNFKFD DYYEIR TQV INEAVNNFKK SSLSGLFDGK IGTIWLIYEF GEIDRAVELF TTHFIEIFEN SQNKNLYSGQ AGILLVGLYF LSKGEID NK LGEEILIRLR EYTLNYIENP ETFCKVGASD VQSNDPYENF GGLLYGHAGV AWLFGEAYKL TGESIYKNGL ELAVDKEL V AYKVDSNNSL QYSQGHRLLP YLATGSAGLL LLINRNKEIL SSKYLKYLTS LERATDVVFC VLPGLFNGFC GLEVANNIY SDIDDNFSGQ KKLIEQLYRY LCVIEEGFVI AGDNGLKITT DIASGFAGVA IGLVSIMDNK LTILPQI

UniProtKB: Protein kinase domain-containing protein

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Macromolecule #2: PneA LP

MacromoleculeName: PneA LP / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Streptococcus pneumoniae (bacteria)
Molecular weightTheoretical: 1.209482 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)

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Macromolecule #3: GUANOSINE-5'-TRIPHOSPHATE

MacromoleculeName: GUANOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 1 / Formula: GTP
Molecular weightTheoretical: 523.18 Da
Chemical component information

ChemComp-GTP:
GUANOSINE-5'-TRIPHOSPHATE / GTP, energy-carrying molecule*YM

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Macromolecule #4: PHOSPHATE ION

MacromoleculeName: PHOSPHATE ION / type: ligand / ID: 4 / Number of copies: 1 / Formula: PO4
Molecular weightTheoretical: 94.971 Da
Chemical component information

ChemComp-PO4:
PHOSPHATE ION

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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

BufferpH: 7.4
Component:
ConcentrationFormulaName
200.0 mMNaClsodium chloride
25.0 mM(HOCH2)3CNH2Tris
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 32.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing #1

Image processing ID1
Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.98 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 204228
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
FSC plot (resolution estimation)

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Image processing #2

Image processing ID2
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.98 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 204228
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
FSC plot (resolution estimation)

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