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- EMDB-20876: Receptor binding protein-focused reconstruction of the Staphyloco... -

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Basic information

Entry
Database: EMDB / ID: EMD-20876
TitleReceptor binding protein-focused reconstruction of the Staphylococcus aureus phage 80alpha baseplateReceptor (biochemistry)
Map dataC6, full particle reconstruction following C6, RBP-focused 3D classification of non-RBP signal-subtracted particles. Locally autosharpened in Phenix and used for final RBP model refinement.
Sample
  • Complex: Staphylococcus phage 80alpha baseplate
    • Protein or peptide: Major Tail Protein
    • Protein or peptide: Distal Tail Protein
    • Protein or peptide: Tail-Associated Lysin
    • Protein or peptide: Receptor Binding ProteinReceptor (biochemistry)
    • Protein or peptide: Fiber Lower
    • Protein or peptide: Fiber Upper
    • Protein or peptide: Tape Measure Protein
Function / homology
Function and homology information


membrane => GO:0016020
Similarity search - Function
Bacteriophage 69, Orf23, major tail protein / Prophage tail endopeptidase / Prophage endopeptidase tail / Siphovirus-type tail component / Phage tail protein RIFT-related domain / BppU, N-terminal / BppU N-terminal domain / Phage major tail protein TP901-1 / Phage tail tube protein / SGNH hydrolase-type esterase domain ...Bacteriophage 69, Orf23, major tail protein / Prophage tail endopeptidase / Prophage endopeptidase tail / Siphovirus-type tail component / Phage tail protein RIFT-related domain / BppU, N-terminal / BppU N-terminal domain / Phage major tail protein TP901-1 / Phage tail tube protein / SGNH hydrolase-type esterase domain / GDSL-like Lipase/Acylhydrolase family / SGNH hydrolase superfamily / Armadillo-type fold
Similarity search - Domain/homology
Major tail protein / Tape measure protein / Uncharacterized protein / Uncharacterized protein / Minor structure protein / BppU_N domain-containing protein / Tail fiber protein
Similarity search - Component
Biological speciesStaphylococcus virus 80alpha
Methodsingle particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsKizziah JL / Dokland T
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious DiseasesR01 AI083255 United States
CitationJournal: PLoS Pathog / Year: 2020
Title: Structure of the host cell recognition and penetration machinery of a Staphylococcus aureus bacteriophage.
Authors: James L Kizziah / Keith A Manning / Altaira D Dearborn / Terje Dokland /
Abstract: Staphylococcus aureus is a common cause of infections in humans. The emergence of virulent, antibiotic-resistant strains of S. aureus is a significant public health concern. Most virulence and ...Staphylococcus aureus is a common cause of infections in humans. The emergence of virulent, antibiotic-resistant strains of S. aureus is a significant public health concern. Most virulence and resistance factors in S. aureus are encoded by mobile genetic elements, and transduction by bacteriophages represents the main mechanism for horizontal gene transfer. The baseplate is a specialized structure at the tip of bacteriophage tails that plays key roles in host recognition, cell wall penetration, and DNA ejection. We have used high-resolution cryo-electron microscopy to determine the structure of the S. aureus bacteriophage 80α baseplate at 3.75 Å resolution, allowing atomic models to be built for most of the major tail and baseplate proteins, including two tail fibers, the receptor binding protein, and part of the tape measure protein. Our structure provides a structural basis for understanding host recognition, cell wall penetration and DNA ejection in viruses infecting Gram-positive bacteria. Comparison to other phages demonstrates the modular design of baseplate proteins, and the adaptations to the host that take place during the evolution of staphylococci and other pathogens.
History
DepositionOct 29, 2019-
Header (metadata) releaseNov 13, 2019-
Map releaseMar 4, 2020-
UpdateMar 4, 2020-
Current statusMar 4, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 4.5
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 4.5
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6v8i
  • Surface level: 4.5
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_20876.png

Downloads & links

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Map

FileDownload / File: emd_20876.map.gz / Format: CCP4 / Size: 512 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationC6, full particle reconstruction following C6, RBP-focused 3D classification of non-RBP signal-subtracted particles. Locally autosharpened in Phenix and used for final RBP model refinement.
Voxel sizeX=Y=Z: 1.21 Å
Density
Contour LevelBy AUTHOR: 4.5 / Movie #1: 4.5
Minimum - Maximum-20.3634 - 27.733955
Average (Standard dev.)-0.0017119119 (±0.98120457)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions512512512
Spacing512512512
CellA=B=C: 619.52 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.211.211.21
M x/y/z512512512
origin x/y/z0.0000.0000.000
length x/y/z619.520619.520619.520
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ512512512
MAP C/R/S321
start NC/NR/NS000
NC/NR/NS512512512
D min/max/mean-20.36327.734-0.002

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Supplemental data

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Sample components

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Entire : Staphylococcus phage 80alpha baseplate

EntireName: Staphylococcus phage 80alpha baseplate
Components
  • Complex: Staphylococcus phage 80alpha baseplate
    • Protein or peptide: Major Tail Protein
    • Protein or peptide: Distal Tail Protein
    • Protein or peptide: Tail-Associated Lysin
    • Protein or peptide: Receptor Binding ProteinReceptor (biochemistry)
    • Protein or peptide: Fiber Lower
    • Protein or peptide: Fiber Upper
    • Protein or peptide: Tape Measure Protein

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Supramolecule #1: Staphylococcus phage 80alpha baseplate

SupramoleculeName: Staphylococcus phage 80alpha baseplate / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: 80alpha baseplate attached to fully formed tails with no capsids
Source (natural)Organism: Staphylococcus virus 80alpha / Strain: ST247
Recombinant expressionOrganism: Staphylococcus aureus (bacteria)
Molecular weightTheoretical: 3.7 MDa

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Macromolecule #1: Major Tail Protein

MacromoleculeName: Major Tail Protein / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus virus 80alpha
Recombinant expressionOrganism: Staphylococcus aureus (bacteria)
SequenceString:
MANMKNSNDR IILFRKAGEK VDATKMLFLT EYGLSHEADT DTEDTMDGSY NTGGSVESTM SGTAKMFYGD DFADEIEDAV VDRVLYEAWE VESRIPGKNG DSAKFKAKYF QGFHNKFELK AEANGIDEYE YEYGVNGRFQ RGFATLPEAV TKKLKATGYR FHDTTKEDAL TSEDLTAIPQ PKVDSSTVTP GEV

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Macromolecule #2: Distal Tail Protein

MacromoleculeName: Distal Tail Protein / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus virus 80alpha
Recombinant expressionOrganism: Staphylococcus aureus (bacteria)
SequenceString: MDIELTKKDG TVIKLSEYGF IVNDIVIDSM QINTKYQDKE NMNGRILMGS NYISRDIVVP CFCKVKNRSD IAYMRDMLYS LTTDIEPMYL REIRRKEELN YRFTQPTSDD YVKLDKNNFP DYEYSRHDQQ IYVNGKQYKV IFNGVINPKQ KGNKVSFELK FETTELPYGE ...String:
MDIELTKKDG TVIKLSEYGF IVNDIVIDSM QINTKYQDKE NMNGRILMGS NYISRDIVVP CFCKVKNRSD IAYMRDMLYS LTTDIEPMYL REIRRKEELN YRFTQPTSDD YVKLDKNNFP DYEYSRHDQQ IYVNGKQYKV IFNGVINPKQ KGNKVSFELK FETTELPYGE SIGTSLELEE NKKVGLWSFD FNIDWHAGGD KRKYTFENLS KGTVYYHGSA PNDQFNMYKK ITIILGEDTE SFVWNLTHAE IMKIEGIKLK AGDRIVYDSF RVYKNGVEIS TETNISQPKF KYGANKFEFN QTVQKVQFDL KFYYK

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Macromolecule #3: Tail-Associated Lysin

MacromoleculeName: Tail-Associated Lysin / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus virus 80alpha
Recombinant expressionOrganism: Staphylococcus aureus (bacteria)
SequenceString: MTITIKPPKG NGAPVPVETT LVKKVNADGV LTFDILENKY TYEVINAIGK RWIVSHVEGE NDKKEYVITV IDRKSEGDRQ LVECTAREIP IDKLMIDRIY VNVTGSFTVE RYFNIVFQGT GMLFEVEGKV KSSKFENGGE GDTRLEMFKK GLEHFGLEYK ITYDKKKDRY ...String:
MTITIKPPKG NGAPVPVETT LVKKVNADGV LTFDILENKY TYEVINAIGK RWIVSHVEGE NDKKEYVITV IDRKSEGDRQ LVECTAREIP IDKLMIDRIY VNVTGSFTVE RYFNIVFQGT GMLFEVEGKV KSSKFENGGE GDTRLEMFKK GLEHFGLEYK ITYDKKKDRY KFVLTPFANQ KASYFISDEV NANAIKLEED ASDFATFIRG YGNYSGEETF EHAGLVMEAR SALAEIYGDI HAEPFKDGKV TDQETMDKEL QSRLKKSLKQ SLSLDFLVLR ESYPEADPQP GDIVQIKSTK LGLNDLVRIV QVKTIRGINN VIVKQDVTLG EFNREQRYMK KVNTAANYVS GLNDVNLSNP SKAAENLKSK VASIAKSTLD LMSRTDLIED KQQKVSSKTV TTSDGTIVHD FIDKSNIKDV KTIGTIGDSV ARGSHAKTNF TEMLGKKLKA KTTNLARGGA TMATVPIGKE AVENSIYRQA EQIRGDLIIL QGTDDDWLHG YWAGVPIGTD KTDTKTFYGA FCSAIEVIRK NNPASKILVM TATRQCPMSG TMIRRKDTDK NKLGLTLEDY VNAQILACSE LDVPVYDAYH TDYFKPYNPA FRKSSMPDGL HPNERGHEVI MYELIKNYYQ FYG

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Macromolecule #4: Receptor Binding Protein

MacromoleculeName: Receptor Binding Protein / type: protein_or_peptide / ID: 4 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus virus 80alpha
Recombinant expressionOrganism: Staphylococcus aureus (bacteria)
SequenceString: MDNKLITDLS RVFDYRYVDE NEYNFKLISD MLTDFNFSLE YHRNKEVFAH NGEQIKYEHL NVTSSVSDFL TYLNGRFSNM VLGHNGDGIN EVKDARVDNT GYDHKTLQDR LYHDYSTLDA FTKKVEKAVD ENYKEYRATE YRFEPKEQEP EFITDLSPYT NAVMQSFWVD ...String:
MDNKLITDLS RVFDYRYVDE NEYNFKLISD MLTDFNFSLE YHRNKEVFAH NGEQIKYEHL NVTSSVSDFL TYLNGRFSNM VLGHNGDGIN EVKDARVDNT GYDHKTLQDR LYHDYSTLDA FTKKVEKAVD ENYKEYRATE YRFEPKEQEP EFITDLSPYT NAVMQSFWVD PRTKIIYMTQ ARPGNHYMLS RLKPNGQFID RLLVKNGGHG THNAYRYIGN ELWIYSAVLD ANENNKFVRF QYRTGEITYG NEMQDVMPNI FNDRYTSAIY NPIENLMIFR REYKASERQL KNSLNFVEVR SADDIDKGID KVLYQMDIPM EYTSDTQPMQ GITYDAGILY WYTGDSKPAN PNYLQGFDIK TKELLFKRRI DIGGVNNNFK GDFQEAEGLD MYYDLETGRK ALLIGVTIGP GNNRHHSIYS IGQRGVNQFL KNIAPQVSMT DSGGRVKPLP IQNPAYLSDI TEVGHYYIYT QDTQNALDFP LPKAFRDAGW FFDVLPGHYN GALRQVLTRN STGRNMLKFE RVIDIFNKKN NGAWNFCPQN AGYWEHIPKS ITKLSDLKIV GLDFYITTEE SNRFTDFPKD FKGIAGWILE VKSNTPGNTT QVLRRNNFPS AHQFLVRNFG TGGVGKWSLF EGKVVE

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Macromolecule #5: Fiber Lower

MacromoleculeName: Fiber Lower / type: protein_or_peptide / ID: 5 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus virus 80alpha
Recombinant expressionOrganism: Staphylococcus aureus (bacteria)
SequenceString: MVVDNFSKDD NLIELQTTSQ YNPVIDTNIS FYESDRGTGV LNFAVTKNNR PLSISSEHVK TFIVLKTDDY NVDRGAYISD ELTIVDAING RLQYVIPNEF LKHSGKVHAQ AFFTQNGSDN VVVERQFSFN IENDLVSGFD GITKLVYIKS IQDTIEAVGK DFNQLKQNMA ...String:
MVVDNFSKDD NLIELQTTSQ YNPVIDTNIS FYESDRGTGV LNFAVTKNNR PLSISSEHVK TFIVLKTDDY NVDRGAYISD ELTIVDAING RLQYVIPNEF LKHSGKVHAQ AFFTQNGSDN VVVERQFSFN IENDLVSGFD GITKLVYIKS IQDTIEAVGK DFNQLKQNMA DTQTLIAKVN DSATKGIQQI EIKQNEAIQA ITATQTSATQ AVTAEFDKIV EKEQAIFERV NEVEQQINGA DLVKGNSTTN WQKSKLTDDY GKAIESYEQS IDSVLSAVNT SRIIHITSAT DAPSFKDIGT VDTPKEDGVD DGSDIPVAPN TLGKSGVLVV YVVDDSTARA TWYPDDSNDE YTKYKISGTW YPFYKKNDGD LTKQFVEETS NNALNQAKQY VDDKFGTTSW QQHKMTEANG QSIQVNLNNA QGDLGYLTAG NYYATRVPDL PGSVESYEGY LSVFVKDDTN KLFNFTPYNS KKIYTRSITN GRLEQQWTVP NEHKSTVLFD GGANGVGTTI NLTEPYTNYS ILLVSGTYPG GVIEGFGLTA LPNAIQLSKA NVVDSDGNGG GIYECLLSKT SSTTLRIDND VYFDLGKTSG SGANANKVTI TKIMGWK

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Macromolecule #6: Fiber Upper

MacromoleculeName: Fiber Upper / type: protein_or_peptide / ID: 6 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus virus 80alpha
Recombinant expressionOrganism: Staphylococcus aureus (bacteria)
SequenceString: MYKIKDVETR IKNDGVDLGD IGCRFYTEDE NTASIRIGIN DKQGRIDLKA HGLTPRLHLF MEDGSIFKNE PLIIDDVVKG FITYKIPKKV IKHAGYVRCK LFLEKEEEKI HVANFSFNIV DSGIESAVAK EIDVKLVDDA ITRILKDNAT DLLSKDFKEK IDKDVISYIE ...String:
MYKIKDVETR IKNDGVDLGD IGCRFYTEDE NTASIRIGIN DKQGRIDLKA HGLTPRLHLF MEDGSIFKNE PLIIDDVVKG FITYKIPKKV IKHAGYVRCK LFLEKEEEKI HVANFSFNIV DSGIESAVAK EIDVKLVDDA ITRILKDNAT DLLSKDFKEK IDKDVISYIE KNESRFKGAK GDKGEPGQPG AKGEAGKKGE QGAPGKNGTV VSINPDTKMW QIDGKDTDIK AEPELLDKIN IANVEGLEDK LQEVEKIKDT TLNDSKTYTD TKIAELVDSA PESMNTLREL AEAIQNNSIS ESVLQQIGSK VSTEDFEEFK QTLNDLYAPK NHNHDERYVL SSQAFTKQQA DSLYQLKSAS QPTVKIWTGT ENEYNYIYQK DPNTLYLIKG

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Macromolecule #7: Tape Measure Protein

MacromoleculeName: Tape Measure Protein / type: protein_or_peptide / ID: 7 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus virus 80alpha
Recombinant expressionOrganism: Staphylococcus aureus (bacteria)
SequenceString: MTEYKIKATI EASVAKFKRQ IDSAVKSVQR FKRVADQTKD VELNANDKKL QKTIKVAKKS LDAFSNKNVK AKLDASIQDL QQKILESNFE LDKLNSKEAS PEVKLQKQKL TKDIAEAENK LSELEKKRVN IDVNADNSKF NRVLKVSKAS LEALNRSKAK AILDVDNSVA ...String:
MTEYKIKATI EASVAKFKRQ IDSAVKSVQR FKRVADQTKD VELNANDKKL QKTIKVAKKS LDAFSNKNVK AKLDASIQDL QQKILESNFE LDKLNSKEAS PEVKLQKQKL TKDIAEAENK LSELEKKRVN IDVNADNSKF NRVLKVSKAS LEALNRSKAK AILDVDNSVA NSKIKRTKEE LKSIPNKTRS RLDVDTRLSI PTIYAFKKSL DALPNKKTTK VDVDTNGLKK VYAYIIKAND NFQRQMGNLA NMFRVFGTVG SNMVGGLLTS SFSILIPVIA SVVPVVFALL NAIKVLTGGV LALGGAVAIA GAGFVAFGAM AISAIKMLND GTLQASSATN EYKKALDGVK SAWTDIIKQN QSAIFTTLAN GLNTVKTAMQ SLQPFFSGIS RGMEEASQSV LKWAENSSVA SRFFNMMNTT GVSVFNKLLS AAGGFGDGLV NVFTQLAPLF QWSADWLDRL GQSFSNWANS AAGENSITRF IEYTKTNLPI IGNIFKNVFA GINNLMNAFS GSSTGIFQSL EQMTAKFREW SEQVGQSQGF KDFVSYIQTN GPLIMQLIGN IARGLVAFAT AMAPIASAVL RVAVAITGWI ANLFEAHPAT AQLVGVIITL VGAFRFLIAP ILAVMDFLGP LAARLVALVT KFGWAKTGTL VLSKAMTSLK GPIKLVTAIF QLLFGKIGLI RNAITGLVTV FGILGGPITI VIGVIAALIA IFVLLWNKNE GFRNFIINAW NAIKTFMVNV WNVLKAVASV VWNAILTAIT TAVSNVYNFI MIVWNQIVAY LQGLWNGIIA IATTVWNLLV TIITTVFTTI MTIVMTIWTA IWTFLSTIWN TIITIATTIW NLLVTVITTV FTTIMTIAMT IWNAIWTFLQ TLWNTIVTVA TKVWNAITTA ISTALQAAWS FISNIWNTIW SFLSGILTTI WNKVVSIFTQ VVSTISDKMS QAWNFIVTKG MQWVSTITST LINFVNRVIQ GFVNVVNKVS QGMTNAVNKI KSFIGDFVSA GADMIRGLIR GIGQMAGQLV DAAKNVAKKA LDAAKSALGI HSPSREFMDV GMYSMLGFVK GIDNHSSKVI RNVSNVADKV VDAFQPTLNA PDISSITGNL SNLGGNINAQ VQHTHSIETS PNMKTVKVEF DVNNDALTSI VNGRNAKRNS EYYL

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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: 8
Component:
ConcentrationNameFormula
10.0 mMtris(hydroxymethyl)aminomethane hydrochloric acid
50.0 mMsodium chlorideNaClSodium chloride
1.0 mMmagnesium sulfateMgSO4
4.0 mMcalcium chlorideCaCl2
GridModel: Quantifoil R2/1 / Material: NICKEL / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV
Details80alpha tails (with baseplates) purified by centrifugation, polyethylene glycol precipitation, and CsCl density gradation

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3.7 µm / Nominal defocus min: 1.7 µm / Nominal magnification: 29000
Sample stageSpecimen holder model: GATAN 626 SINGLE TILT LIQUID NITROGEN CRYO TRANSFER HOLDER
Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: DIRECT ELECTRON DE-20 (5k x 3k) / Digitization - Frames/image: 1-37 / Number grids imaged: 10 / Number real images: 6483 / Average exposure time: 1.156 sec. / Average electron dose: 104.33 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 515106
CTF correctionSoftware - Name: Gctf (ver. 1.06_sm_30_cu8.0_x86_64)
Startup modelType of model: OTHER / Details: EMAN2.0 initial model generation
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 2.0)
Final 3D classificationSoftware - Name: RELION (ver. 3.0-beta-2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0-beta-2)
Final reconstructionApplied symmetry - Point group: C6 (6 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0-beta-2) / Number images used: 66861
FSC plot (resolution estimation)

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