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- EMDB-44345: Cryo-EM structure of E. coli cellulose synthase subunit C with ce... -

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Basic information

Entry
Database: EMDB / ID: EMD-44345
TitleCryo-EM structure of E. coli cellulose synthase subunit C with cellotetraose
Map dataBcsC_cellotetraose_sharpened map
Sample
  • Complex: Bacterial cellulose synthase subunit C in nanodisc with cellotetraose
    • Protein or peptide: Cellulose synthase operon protein C
KeywordsPorin / BcsC outer membrane protein / Tetra-tricopeptide (TPR) repeats / cellooligosaccharide / Membrane Protein
Function / homology
Function and homology information


cellulose biosynthetic process / cell outer membrane
Similarity search - Function
Cellulose synthase operon C, C-terminal / Cellulose synthase operon protein C C-terminus (BCSC_C) / : / Tetratricopeptide repeat / Tetratricopeptide repeat / TPR repeat region circular profile. / TPR repeat profile. / Tetratricopeptide repeats / Tetratricopeptide repeat / Tetratricopeptide-like helical domain superfamily
Similarity search - Domain/homology
Cellulose synthase operon protein C
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsVerma P / Zimmer J
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM144130 United States
CitationJournal: Nat Commun / Year: 2024
Title: Insights into phosphoethanolamine cellulose synthesis and secretion across the Gram-negative cell envelope.
Authors: Preeti Verma / Ruoya Ho / Schuyler A Chambers / Lynette Cegelski / Jochen Zimmer /
Abstract: Phosphoethanolamine (pEtN) cellulose is a naturally occurring modified cellulose produced by several Enterobacteriaceae. The minimal components of the E. coli cellulose synthase complex include the ...Phosphoethanolamine (pEtN) cellulose is a naturally occurring modified cellulose produced by several Enterobacteriaceae. The minimal components of the E. coli cellulose synthase complex include the catalytically active BcsA enzyme, a hexameric semicircle of the periplasmic BcsB protein, and the outer membrane (OM)-integrated BcsC subunit containing periplasmic tetratricopeptide repeats (TPR). Additional subunits include BcsG, a membrane-anchored periplasmic pEtN transferase associated with BcsA, and BcsZ, a periplasmic cellulase of unknown biological function. While cellulose synthesis and translocation by BcsA are well described, little is known about its pEtN modification and translocation across the cell envelope. We show that the N-terminal cytosolic domain of BcsA positions three BcsG copies near the nascent cellulose polymer. Further, the semicircle's terminal BcsB subunit tethers the N-terminus of a single BcsC protein in a trans-envelope secretion system. BcsC's TPR motifs bind a putative cello-oligosaccharide near the entrance to its OM pore. Additionally, we show that only the hydrolytic activity of BcsZ but not the subunit itself is necessary for cellulose secretion, suggesting a secretion mechanism based on enzymatic removal of translocation incompetent cellulose. Lastly, protein engineering introduces cellulose pEtN modification in orthogonal cellulose biosynthetic systems. These findings advance our understanding of pEtN cellulose modification and secretion.
History
DepositionMar 30, 2024-
Header (metadata) releaseFeb 19, 2025-
Map releaseFeb 19, 2025-
UpdateMay 28, 2025-
Current statusMay 28, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_44345.png

Downloads & links

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Map

FileDownload / File: emd_44345.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationBcsC_cellotetraose_sharpened map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.08 Å/pix.
x 360 pix.
= 388.8 Å		1.08 Å/pix.
x 360 pix.
= 388.8 Å		1.08 Å/pix.
x 360 pix.
= 388.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.08 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.39
Minimum - Maximum-1.5744195 - 2.6899855
Average (Standard dev.)-0.00066021783 (±0.031064004)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 388.80002 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: BcsC cellotetraose half map A

Fileemd_44345_half_map_1.map
AnnotationBcsC_cellotetraose_half map_A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: BcsC cellotetraose half map B

Fileemd_44345_half_map_2.map
AnnotationBcsC_cellotetraose_half map_B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Bacterial cellulose synthase subunit C in nanodisc with cellotetraose

EntireName: Bacterial cellulose synthase subunit C in nanodisc with cellotetraose
Components
  • Complex: Bacterial cellulose synthase subunit C in nanodisc with cellotetraose
    • Protein or peptide: Cellulose synthase operon protein C

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Supramolecule #1: Bacterial cellulose synthase subunit C in nanodisc with cellotetraose

SupramoleculeName: Bacterial cellulose synthase subunit C in nanodisc with cellotetraose
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Protein: Bacterial cellulose synthase subunit C (BcsC) Ligand: Cellotetraose
Source (natural)Organism: Escherichia coli (E. coli)

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Macromolecule #1: Cellulose synthase operon protein C

MacromoleculeName: Cellulose synthase operon protein C / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 126.827016 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MHHHHHHHHV EAAPTAQQQL LEQVRLGEAT HREDLVQQSL YRLELIDPNN PDVVAARFRS LLRQGDIDGA QKQLDRLSQL APSSNAYKS SRTTMLLSTP DGRQALQQAR LQATTGHAEE AVASYNKLFN GAPPEGDIAV EYWSTVAKIP ARRGEAINQL K RINADAPG ...String:
MHHHHHHHHV EAAPTAQQQL LEQVRLGEAT HREDLVQQSL YRLELIDPNN PDVVAARFRS LLRQGDIDGA QKQLDRLSQL APSSNAYKS SRTTMLLSTP DGRQALQQAR LQATTGHAEE AVASYNKLFN GAPPEGDIAV EYWSTVAKIP ARRGEAINQL K RINADAPG NTGLQNNLAL LLFSSDRRDE GFAVLEQMAK SNAGREGASK IWYGQIKDMP VSDASVSALK KYLSIFSDGD SV AAAQSQL AEQQKQLADP AFRARAQGLA AVDSGMAGKA IPELQQAVRA NPKDSEALGA LGQAYSQKGD RANAVANLEK ALA LDPHSS NNDKWNSLLK VNRYWLAIQQ GDAALKANNP DRAERLFQQA RNVDNTDSYA VLGLGDVAMA RKDYPAAERY YQQT LRMDS GNTNAVRGLA NIYRQQSPEK AEAFIASLSA SQRRSIDDIE RSLQNDRLAQ QAEALENQGK WAQAAALQRQ RLALD PGSV WITYRLSQDL WQAGQRSQAD TLMRNLAQQK SNDPEQVYAY GLYLSGHDQD RAALAHINSL PRAQWNSNIQ ELVNRL QSD QVLETANRLR ESGKEAEAEA MLRQQPPSTR IDLTLADWAQ QRRDYTAARA AYQNVLTREP ANADAILGLT EVDIAAG DK AAARSQLAKL PATDNASLNT QRRVALAQAQ LGDTAAAQRT FNKLIPQAKS QPPSMESAMV LRDGAKFEAQ AGDPTQAL E TYKDAMVASG VTTTRPQDND TFTRLTRNDE KDDWLKRGVR SDAADLYRQQ DLNVTLEHDY WGSSGTGGYS DLKAHTTML QVDAPYSDGR MFFRSDFVNM NVGSFSTNAD GKWDDNWGTC TLQDCSGNRS QSDSGASVAV GWRNDVWSWD IGTTPMGFNV VDVVGGISY SDDIGPLGYT VNAHRRPISS SLLAFGGQKD SPSNTGKKWG GVRADGVGLS LSYDKGEANG VWASLSGDQL T GKNVEDNW RVRWMTGYYY KVINQNNRRV TIGLNNMIWH YDKDLSGYSL GQGGYYSPQE YLSFAIPVMW RERTENWSWE LG ASGSWSH SRTKTMPRYP LMNLIPTDWQ EEAARQSNDG GSSQGFGYTA RALLERRVTS NWFVGTAIDI QQAKDYAPSH FLL YVRYSA AGWQGDMDLP PQPLIPYADW

UniProtKB: Cellulose synthase operon protein C

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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: 8.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.4 µm / Nominal defocus min: 1.2 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: INSILICO MODEL
In silico model: AlphaFold2 predicted BcsC model(AF-P37650-F1-model_v4)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 60119
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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