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- EMDB-28660: Cryo-EM structure of S. aureus BlaR1 TM and zinc metalloprotease ... -
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Open data
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Basic information
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Title | Cryo-EM structure of S. aureus BlaR1 TM and zinc metalloprotease domain | |||||||||
![]() | Sharpened map from cryosparc local refinement | |||||||||
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![]() | Antibiotic resistance / beta-lactam antibiotics / MRSA / BlaR1 / MecR1 / cryo-EM / transmembrane signalling / SIGNALING PROTEIN | |||||||||
Function / homology | ![]() | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.8 Å | |||||||||
![]() | Worrall LJ / Alexander JAN / Vuckovic M / Strynadka NCJ | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural basis of broad-spectrum β-lactam resistance in Staphylococcus aureus. Authors: J Andrew N Alexander / Liam J Worrall / Jinhong Hu / Marija Vuckovic / Nidhi Satishkumar / Raymond Poon / Solmaz Sobhanifar / Federico I Rosell / Joshua Jenkins / Daniel Chiang / Wesley A ...Authors: J Andrew N Alexander / Liam J Worrall / Jinhong Hu / Marija Vuckovic / Nidhi Satishkumar / Raymond Poon / Solmaz Sobhanifar / Federico I Rosell / Joshua Jenkins / Daniel Chiang / Wesley A Mosimann / Henry F Chambers / Mark Paetzel / Som S Chatterjee / Natalie C J Strynadka / ![]() ![]() Abstract: Broad-spectrum β-lactam antibiotic resistance in Staphylococcus aureus is a global healthcare burden. In clinical strains, resistance is largely controlled by BlaR1, a receptor that senses β- ...Broad-spectrum β-lactam antibiotic resistance in Staphylococcus aureus is a global healthcare burden. In clinical strains, resistance is largely controlled by BlaR1, a receptor that senses β-lactams through the acylation of its sensor domain, inducing transmembrane signalling and activation of the cytoplasmic-facing metalloprotease domain. The metalloprotease domain has a role in BlaI derepression, inducing blaZ (β-lactamase PC1) and mecA (β-lactam-resistant cell-wall transpeptidase PBP2a) expression. Here, overcoming hurdles in isolation, we show that BlaR1 cleaves BlaI directly, as necessary for inactivation, with no requirement for additional components as suggested previously. Cryo-electron microscopy structures of BlaR1-the wild type and an autocleavage-deficient F284A mutant, with or without β-lactam-reveal a domain-swapped dimer that we suggest is critical to the stabilization of the signalling loops within. BlaR1 undergoes spontaneous autocleavage in cis between Ser283 and Phe284 and we describe the catalytic mechanism and specificity underlying the self and BlaI cleavage. The structures suggest that allosteric signalling emanates from β-lactam-induced exclusion of the prominent extracellular loop bound competitively in the sensor-domain active site, driving subsequent dynamic motions, including a shift in the sensor towards the membrane and accompanying changes in the zinc metalloprotease domain. We propose that this enhances the expulsion of autocleaved products from the active site, shifting the equilibrium to a state that is permissive of efficient BlaI cleavage. Collectively, this study provides a structure of a two-component signalling receptor that mediates action-in this case, antibiotic resistance-through the direct cleavage of a repressor. | |||||||||
History |
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Structure visualization
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 97 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 17.7 KB 17.7 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 9.9 KB | Display | ![]() |
Images | ![]() | 114.2 KB | ||
Filedesc metadata | ![]() | 6.2 KB | ||
Others | ![]() ![]() | 95.7 MB 95.7 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 8exrMC ![]() 8expC ![]() 8exqC ![]() 8exsC ![]() 8extC C: citing same article ( M: atomic model generated by this map |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
EMDB pages | ![]() ![]() |
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Map
File | ![]() | ||||||||||||||||||||
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Annotation | Sharpened map from cryosparc local refinement | ||||||||||||||||||||
Voxel size | X=Y=Z: 0.77 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: Half map from cryosparc refinement. Maps have a...
File | emd_28660_half_map_1.map | ||||||||||||
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Annotation | Half map from cryosparc refinement. Maps have a lot of high resolution noise. Lowpass filtering to 3.8 Ang. (FSC resolution of final map) reveals structure. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map from cryosparc refinement. Maps have a...
File | emd_28660_half_map_2.map | ||||||||||||
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Annotation | Half map from cryosparc refinement. Maps have a lot of high resolution noise. Lowpass filtering to 3.8 Ang. (FSC resolution of final map) reveals structure. | ||||||||||||
Projections & Slices |
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Density Histograms |
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Sample components
-Entire : Dimeric complex of S. aureus BlaR1
Entire | Name: Dimeric complex of S. aureus BlaR1 |
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Components |
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-Supramolecule #1: Dimeric complex of S. aureus BlaR1
Supramolecule | Name: Dimeric complex of S. aureus BlaR1 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 142 KDa |
-Macromolecule #1: Beta-lactam sensor/signal transducer BlaR1
Macromolecule | Name: Beta-lactam sensor/signal transducer BlaR1 / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 71.269703 KDa |
Recombinant expression | Organism: ![]() |
Sequence | String: MAKLLIMSIV SFCFIFLLLL FFRYILKRYF NYMLNYKVWY LTLLAGLIPF IPIKFSLFKF NNVNNQAPTV ESKSHDLNHN INTTKPIQE FATDIHKFNW DSIDNICTVI WIVLVIILSF KFLKALLYLK YLKKQSLYLN ENEKNKIDTI LFNHQYKKNI V IRKAETIQ ...String: MAKLLIMSIV SFCFIFLLLL FFRYILKRYF NYMLNYKVWY LTLLAGLIPF IPIKFSLFKF NNVNNQAPTV ESKSHDLNHN INTTKPIQE FATDIHKFNW DSIDNICTVI WIVLVIILSF KFLKALLYLK YLKKQSLYLN ENEKNKIDTI LFNHQYKKNI V IRKAETIQ SPITFWYGKY IILIPSSYFK SVIDKRLKYI ILHEYAHAKN RDTLHLIIFN IFSIIMSYNP LVHIVKRKII HD NEVEADR FVLNNINKNE FKTYAESIMD SVLNVPFFNK NILSHSFNGK KSLLKRRLIN IKEANLKKQS KLILIFICIF TFL LMVIQS QFLMGQSITD YNYKKPLHND YQILDKSKIF GSNSGSFVMY SMKKDKYYIY NEKESRKRYS PNSTYKIYLA MFGL DRHII NDENSRMSWN HKHYPFDAWN KEQDLNTAMQ NSVNWYFERI SDQIPKNYTA TQLKQLNYGN KNLGSYKSYW MEDSL KISN LEQVIVFKNM MEQNNHFSKK AKNQLSSSLL IKKNEKYELY GKTGTGIVNG KYNNGWFVGY VITNHDKYYF ATHLSD GKP SGKNAELISE KILKEMGVLN GQELALVPRG SSAHHHHHH UniProtKB: BlaR1 |
-Macromolecule #2: ZINC ION
Macromolecule | Name: ZINC ION / type: ligand / ID: 2 / Number of copies: 2 / Formula: ZN |
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Molecular weight | Theoretical: 65.409 Da |
-Macromolecule #3: (2S)-3-{[{[(2S)-2,3-DIHYDROXYPROPYL]OXY}(HYDROXY)PHOSPHORYL]OXY}-...
Macromolecule | Name: (2S)-3-{[{[(2S)-2,3-DIHYDROXYPROPYL]OXY}(HYDROXY)PHOSPHORYL]OXY}-2-[(6E)-HEXADEC-6-ENOYLOXY]PROPYL (8E)-OCTADEC-8-ENOATE type: ligand / ID: 3 / Number of copies: 2 / Formula: P6L |
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Molecular weight | Theoretical: 746.991 Da |
Chemical component information | ![]() ChemComp-P6L: |
-Macromolecule #4: PHOSPHATE ION
Macromolecule | Name: PHOSPHATE ION / type: ligand / ID: 4 / Number of copies: 3 / Formula: PO4 |
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Molecular weight | Theoretical: 94.971 Da |
Chemical component information | ![]() ChemComp-PO4: |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.5 mg/mL |
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Buffer | pH: 7.5 / Details: 20 mM HEPES, pH 7.5, 150 mM sodium chloride |
Vitrification | Cryogen name: ETHANE |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 50.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.5 µm |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |