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- EMDB-50591: Composite map of the mycobacterial PafBC-bound transcription init... -

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Basic information

Entry
Database: EMDB / ID: EMD-50591
TitleComposite map of the mycobacterial PafBC-bound transcription initiation complex
Map dataSharpened map.
Sample
  • Complex: DNA-directed RNA polymerase with transcriptional activator PafBC
    • Protein or peptide: x 8 types
    • DNA: x 2 types
  • Ligand: x 2 types
KeywordsRNAP / PafBC / sigma adaptation / WYL domain / TRANSCRIPTION
Function / homology
Function and homology information


sigma factor activity / bacterial-type RNA polymerase core enzyme binding / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed RNA polymerase / DNA-directed RNA polymerase activity / protein dimerization activity / response to antibiotic / DNA-templated transcription ...sigma factor activity / bacterial-type RNA polymerase core enzyme binding / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed RNA polymerase / DNA-directed RNA polymerase activity / protein dimerization activity / response to antibiotic / DNA-templated transcription / positive regulation of DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / cytoplasm
Similarity search - Function
PafC, HTH domain / PafC helix-turn-helix domain / Protein PafC / : / WYL domain profile. / WYL domain / WYL domain / RNA polymerase-binding protein RbpA / RbpA superfamily / RNA polymerase-binding protein ...PafC, HTH domain / PafC helix-turn-helix domain / Protein PafC / : / WYL domain profile. / WYL domain / WYL domain / RNA polymerase-binding protein RbpA / RbpA superfamily / RNA polymerase-binding protein / Sigma-70 factors family signature 1. / RNA polymerase sigma factor RpoD, C-terminal / RNA polymerase sigma factor RpoD / : / RNA polymerase sigma-70 region 1.2 / Sigma-70 factor, region 1.2 / RNA polymerase sigma-70 region 3 / Sigma-70 region 3 / Sigma-70 factors family signature 2. / RNA polymerase sigma-70 / RNA polymerase sigma-70 region 4 / Sigma-70, region 4 / RNA polymerase sigma-70 region 2 / RNA polymerase sigma-70 like domain / Sigma-70 region 2 / RNA polymerase sigma factor, region 2 / RNA polymerase sigma factor, region 3/4-like / DNA-directed RNA polymerase, omega subunit / DNA-directed RNA polymerase, subunit beta-prime, bacterial type / DNA-directed RNA polymerase, beta subunit, external 1 domain superfamily / DNA-directed RNA polymerase, beta subunit, external 1 domain / RNA polymerase beta subunit external 1 domain / RNA polymerase, alpha subunit, C-terminal / Bacterial RNA polymerase, alpha chain C terminal domain / DNA-directed RNA polymerase beta subunit, bacterial-type / DNA-directed RNA polymerase, alpha subunit / DNA-directed RNA polymerase, subunit beta-prime / RNA polymerase Rpb6 / RNA polymerase, subunit omega/Rpo6/RPB6 / RNA polymerase Rpb6 / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase Rpb1, domain 3 superfamily / RPB6/omega subunit-like superfamily / DNA-directed RNA polymerase, insert domain / DNA-directed RNA polymerase, RpoA/D/Rpb3-type / RNA polymerase Rpb3/RpoA insert domain / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerases D / RNA polymerase Rpb1, clamp domain superfamily / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 1 / RNA polymerase, beta subunit, protrusion / RNA polymerase beta subunit / RNA polymerase, alpha subunit / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / RNA polymerase Rpb1, domain 2 / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase, N-terminal / RNA polymerase Rpb2, domain 2 / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase Rpb2, domain 2 / RNA polymerase I subunit A N-terminus / RNA polymerase, RBP11-like subunit / RNA polymerase, beta subunit, conserved site / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 3 / RNA polymerase Rpb2, OB-fold / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 3 / RNA polymerases beta chain signature. / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain / DNA-directed RNA polymerase, subunit 2 / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain superfamily / RNA polymerase Rpb2, domain 6 / Winged helix-like DNA-binding domain superfamily
Similarity search - Domain/homology
DNA-directed RNA polymerase subunit beta' / DNA-directed RNA polymerase subunit alpha / RNA polymerase sigma factor SigA / DNA-directed RNA polymerase subunit omega / RNA polymerase-binding protein RbpA / Protein pafC / Transcriptional regulator-like protein / DNA-directed RNA polymerase subunit beta
Similarity search - Component
Biological speciesMycolicibacterium smegmatis MC2 155 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.0 Å
AuthorsZdanowicz R / Schilling CM / Rabl J / Mueller AU / Boehringer D / Glockshuber R / Weber-Ban E
Funding support Switzerland, 1 items
OrganizationGrant numberCountry
Swiss National Science Foundation310030_215606 Switzerland
CitationJournal: Sci Adv / Year: 2025
Title: Single-stranded DNA binding to the transcription factor PafBC triggers the mycobacterial DNA damage response.
Authors: Charlotte M Schilling / Rafal Zdanowicz / Julius Rabl / Andreas U Müller / Daniel Boehringer / Rudi Glockshuber / Eilika Weber-Ban /
Abstract: The DNA damage response in mycobacteria is controlled by the heterodimeric transcription factor PafBC, a member of the WYL domain-containing protein family. It has been shown that PafBC induces ...The DNA damage response in mycobacteria is controlled by the heterodimeric transcription factor PafBC, a member of the WYL domain-containing protein family. It has been shown that PafBC induces transcription of its regulon by reprogramming the housekeeping RNA polymerase holoenzyme to recognize PafBC-dependent promoters through sigma adaptation. However, the mechanism by which DNA damage is sensed and translated into PafBC activation has remained unclear. Here, we demonstrate that the binding of single-stranded DNA (ssDNA) to the WYL domains of PafBC activates the transcription factor. Our cryo-electron microscopy structure of full-length PafBC in its active conformation, bound to the transcription initiation complex, reveals a previously unknown mode of interaction between the ssDNA and the WYL domains. Using biochemical experiments, we show that short ssDNA fragments bind to PafBC dynamically, resulting in deactivation as ssDNA levels decrease postrepair. Our findings shed light on the mechanism linking DNA damage to PafBC activation and expand our understanding of WYL domain-containing proteins.
History
DepositionJun 10, 2024-
Header (metadata) releaseFeb 12, 2025-
Map releaseFeb 12, 2025-
UpdateJul 2, 2025-
Current statusJul 2, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_50591.png

Downloads & links

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Map

FileDownload / File: emd_50591.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharpened map.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
1.12 Å/pix.
x 360 pix.
= 403.2 Å		1.12 Å/pix.
x 360 pix.
= 403.2 Å		1.12 Å/pix.
x 360 pix.
= 403.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.12 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 3.3
Minimum - Maximum-48.434820000000002 - 66.076130000000006
Average (Standard dev.)-0.000000000000418 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 403.2 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Unsharpened map.

Fileemd_50591_additional_1.map
AnnotationUnsharpened map.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : DNA-directed RNA polymerase with transcriptional activator PafBC

EntireName: DNA-directed RNA polymerase with transcriptional activator PafBC
Components
  • Complex: DNA-directed RNA polymerase with transcriptional activator PafBC
    • Protein or peptide: DNA-directed RNA polymerase subunit alpha
    • Protein or peptide: DNA-directed RNA polymerase subunit beta
    • Protein or peptide: DNA-directed RNA polymerase subunit beta'
    • Protein or peptide: DNA-directed RNA polymerase subunit omega
    • Protein or peptide: RNA polymerase sigma factor SigA
    • Protein or peptide: RNA polymerase-binding protein RbpA
    • DNA: recA-op non-template strand
    • DNA: recA-op template strand
    • Protein or peptide: PafC
    • Protein or peptide: Transcriptional regulator-like protein
  • Ligand: ZINC ION
  • Ligand: MAGNESIUM ION

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Supramolecule #1: DNA-directed RNA polymerase with transcriptional activator PafBC

SupramoleculeName: DNA-directed RNA polymerase with transcriptional activator PafBC
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#10
Source (natural)Organism: Mycolicibacterium smegmatis MC2 155 (bacteria)

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Macromolecule #1: DNA-directed RNA polymerase subunit alpha

MacromoleculeName: DNA-directed RNA polymerase subunit alpha / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Mycolicibacterium smegmatis MC2 155 (bacteria)
Molecular weightTheoretical: 37.959441 KDa
SequenceString: MLISQRPTLS EETVAENRSR FVIEPLEPGF GYTLGNSLRR TLLSSIPGAA VTSIRIDGVL HEFTTVPGVK EDVTDIILNL KGLVVSSDD DEPVTMYLRK QGPGVVTAGD IVPPAGVTVH NPDMHIATLN DKGKLEVELV VERGRGYVPA VQNKASGAEI G RIPVDSIY ...String:
MLISQRPTLS EETVAENRSR FVIEPLEPGF GYTLGNSLRR TLLSSIPGAA VTSIRIDGVL HEFTTVPGVK EDVTDIILNL KGLVVSSDD DEPVTMYLRK QGPGVVTAGD IVPPAGVTVH NPDMHIATLN DKGKLEVELV VERGRGYVPA VQNKASGAEI G RIPVDSIY SPVLKVTYKV EATRVEQRTD FDKLIIDVET KNSISPRDAL ASAGGTLVEL FGLARELNAD SEHIEIGPSP AE ADHIASF ALPIDDLDLT VRSYNCLKRE GVHTVGELVA RTESDLLDIR NFGQKSIDEV KIKLHQLGLS LKDSPATFDP SEV AGYDAA TGTWTSDAGY DLDDNQDYAE TEQL

UniProtKB: DNA-directed RNA polymerase subunit alpha

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Macromolecule #2: DNA-directed RNA polymerase subunit beta

MacromoleculeName: DNA-directed RNA polymerase subunit beta / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Mycolicibacterium smegmatis MC2 155 (bacteria)
Molecular weightTheoretical: 128.680141 KDa
SequenceString: MLEGCILAVS SQSKSNAITN NSVPGAPNRV SFAKLREPLE VPGLLDVQTD SFEWLVGSDR WRQAAIDRGE ENPVGGLEEV LAELSPIED FSGSMSLSFS DPRFDEVKAS VDECKDKDMT YAAPLFVTAE FINNNTGEIK SQTVFMGDFP MMTEKGTFII N GTERVVVS ...String:
MLEGCILAVS SQSKSNAITN NSVPGAPNRV SFAKLREPLE VPGLLDVQTD SFEWLVGSDR WRQAAIDRGE ENPVGGLEEV LAELSPIED FSGSMSLSFS DPRFDEVKAS VDECKDKDMT YAAPLFVTAE FINNNTGEIK SQTVFMGDFP MMTEKGTFII N GTERVVVS QLVRSPGVYF DETIDKSTEK TLHSVKVIPG RGAWLEFDVD KRDTVGVRID RKRRQPVTVL LKALGWTNEQ IV ERFGFSE IMMGTLEKDT TSGTDEALLD IYRKLRPGEP PTKESAQTLL ENLFFKEKRY DLARVGRYKV NKKLGLNAGK PIT SSTLTE EDVVATIEYL VRLHEGQTSM TVPGGVEVPV EVDDIDHFGN RRLRTVGELI QNQIRVGLSR MERVVRERMT TQDV EAITP QTLINIRPVV AAIKEFFGTS QLSQFMDQNN PLSGLTHKRR LSALGPGGLS RERAGLEVRD VHPSHYGRMC PIETP EGPN IGLIGSLSVY ARVNPFGFIE TPYRKVENGV VTDQIDYLTA DEEDRHVVAQ ANSPTDENGR FTEDRVMVRK KGGEVE FVS ADQVDYMDVS PRQMVSVATA MIPFLEHDDA NRALMGANMQ RQAVPLVRSE APLVGTGMEL RAAIDAGDVV VADKTGV IE EVSADYITVM ADDGTRQSYR LRKFARSNHG TCANQRPIVD AGQRVEAGQV IADGPCTQNG EMALGKNLLV AIMPWEGH N YEDAIILSNR LVEEDVLTSI HIEEHEIDAR DTKLGAEEIT RDIPNVSDEV LADLDERGIV RIGAEVRDGD ILVGKVTPK GETELTPEER LLRAIFGEKA REVRDTSLKV PHGESGKVIG IRVFSREDDD ELPAGVNELV RVYVAQKRKI SDGDKLAGRH GNKGVIGKI LPVEDMPFLP DGTPVDIILN THGVPRRMNI GQILETHLGW VAKAGWNIDV AAGVPDWASK LPEELYSAPA D STVATPVF DGAQEGELAG LLGSTLPNRD GEVMVDADGK STLFDGRSGE PFPYPVTVGY MYILKLHHLV DDKIHARSTG PY SMITQQP LGGKAQFGGQ RFGEMECWAM QAYGAAYTLQ ELLTIKSDDT VGRVKVYEAI VKGENIPEPG IPESFKVLLK ELQ SLCLNV EVLSSDGAAI EMRDGDDEDL ERAAANLGIN LSRNESASVE DLA

UniProtKB: DNA-directed RNA polymerase subunit beta

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Macromolecule #3: DNA-directed RNA polymerase subunit beta'

MacromoleculeName: DNA-directed RNA polymerase subunit beta' / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Mycolicibacterium smegmatis MC2 155 (bacteria)
Molecular weightTheoretical: 146.712891 KDa
SequenceString: MLDVNFFDEL RIGLATADDI RNWSYGEVKK PETINYRTLK PEKDGLFCEK IFGPTRDWEC YCGKYKRVRF KGIICERCGV EVTRAKVRR ERMGHIELAA PVTHIWYFKG VPSRLGYLLD LAPKDLEKII YFAAYVITSV DDEMRHNELS TLEAEMAVEK K AVEDQRDA ...String:
MLDVNFFDEL RIGLATADDI RNWSYGEVKK PETINYRTLK PEKDGLFCEK IFGPTRDWEC YCGKYKRVRF KGIICERCGV EVTRAKVRR ERMGHIELAA PVTHIWYFKG VPSRLGYLLD LAPKDLEKII YFAAYVITSV DDEMRHNELS TLEAEMAVEK K AVEDQRDA DLEARAQKLE ADLAELEAEG AKSDVRRKVR DSGEREMRQL RDRAQRELDR LDEIWNTFTK LAPKQLIVDE VL YRELQDR YGEYFTGAMG AESIKKLIEN FDIDAEAESL REVIRSGKGQ KKLRALKRLK VVAAFQQSGN SPMGMVLDAV PVI PPELRP MVQLDGGRFA TSDLNDLYRR VINRNNRLKR LIDLGAPEII VNNEKRMLQE SVDALFDNGR RGRPVTGPGN RPLK SLSDL LKGKQGRFRQ NLLGKRVDYS GRSVIVVGPQ LKLHQCGLPK LMALELFKPF VMKRLVDLNH AQNIKSAKRM VERQR PQVW DVLEEVIAEH PVLLNRAPTL HRLGIQAFEP QLVEGKAIQL HPLVCEAFNA DFDGDQMAVH LPLSAEAQAE ARILML SSN NILSPASGKP LAMPRLDMVT GLYYLTTLVE GATGEYQAAT KDAPEQGVYS SPAEAIMAMD RGALSVRAKI KVRLTEL RP PTDLEAQLFE NGWKPGDAWT AETTLGRVMF NELLPKSYPF VNEQMHKKVQ ARIINDLAER FPMIVVAQTV DKLKDAGF Y WATRSGVTVS MADVLVPPQK QEILERHEAE ADAIERKYQR GALNHTERNE SLVKIWQDAT EEVGKALEEF YPADNPIIT IVKSGATGNL TQTRTLAGMK GLVTNPKGEF IPRPIKSSFR EGLTVLEYFI NTHGARKGLA DTALRTADSG YLTRRLVDVS QDVIVREHD CETERGINVT LAERGPDGTL IRDAHVETSA FARTLATDAV DANGNVIIER GHDLGDPAID ALLAAGITTV K VRSVLTCT SATGVCAMCY GRSMATGKLV DIGEAVGIVA AQSIGEPGTQ LTMRTFHQGG VTGGADIVGG LPRVQELFEA RV PRNKAPI ADVAGRVRLE ESDKFFKITI VPDDGGEEVV YDKLSKRQRL RVITHEDGTE GVLSDGDHVE VGDQLMEGAA DPH EVLRVQ GPREVQIHLV KEVQEVYRAQ GVSIHDKHIE VIVRQMLRRV TIIDSGSTEF LPGSLTERAE FEAENRRVVA EGGE PAAGR PVLMGITKAS LATDSWLSAA SFQETTRVLT DAAINCRSDK LNGLKENVII GKLIPAGTGI SRYRNIQVQP TEEAR AAAY TIPSYEDQYY SPDFGQATGA AVPLDDYGYS DYR

UniProtKB: DNA-directed RNA polymerase subunit beta'

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Macromolecule #4: DNA-directed RNA polymerase subunit omega

MacromoleculeName: DNA-directed RNA polymerase subunit omega / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Mycolicibacterium smegmatis MC2 155 (bacteria)
Molecular weightTheoretical: 11.544763 KDa
SequenceString:
MSTPHADAQL NAADDLGIDS SAASAYDTPL GITNPPIDEL LSRASSKYAL VIYAAKRARQ INDYYNQLGD GILEYVGPLV EPGLQEKPL SIALREIHGD LLEHTEGE

UniProtKB: DNA-directed RNA polymerase subunit omega

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Macromolecule #5: RNA polymerase sigma factor SigA

MacromoleculeName: RNA polymerase sigma factor SigA / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycolicibacterium smegmatis MC2 155 (bacteria)
Molecular weightTheoretical: 51.573551 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MAATKASPAT EEPVKRTATK TPAKKAPAKR AAKSAAAKAG GKAPAKKAPA KRAAKGTAAK PEDGVTDDLE VTDDLEAEPG EDLDVEDTD LELDDLDSDD DTAVEDEEEE ADAATPAVAT AKAADDDIDE PSEKDKASGD FVWDEEESEA LRQARKDAEL T ASADSVRA ...String:
MAATKASPAT EEPVKRTATK TPAKKAPAKR AAKSAAAKAG GKAPAKKAPA KRAAKGTAAK PEDGVTDDLE VTDDLEAEPG EDLDVEDTD LELDDLDSDD DTAVEDEEEE ADAATPAVAT AKAADDDIDE PSEKDKASGD FVWDEEESEA LRQARKDAEL T ASADSVRA YLKQIGKVAL LNAEEEVELA KRIEAGLYAT QKLAELAEKG EKLPVQQRRD MQWICRDGDR AKNHLLEANL RL VVSLAKR YTGRGMAFLD LIQEGNLGLI RAVEKFDYTK GYKFSTYATW WIRQAITRAM ADQARTIRIP VHMVEVINKL GRI QRELLQ DLGREPTPEE LAKEMDITPE KVLEIQQYAR EPISLDQTIG DEGDSQLGDF IEDSEAVVAV DAVSFTLLQD QLQS VLETL SEREAGVVRL RFGLTDGQPR TLDEIGQVYG VTRERIRQIE SKTMSKLRHP SRSQVLRDYL D

UniProtKB: RNA polymerase sigma factor SigA

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Macromolecule #6: RNA polymerase-binding protein RbpA

MacromoleculeName: RNA polymerase-binding protein RbpA / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycolicibacterium smegmatis MC2 155 (bacteria)
Molecular weightTheoretical: 13.078731 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MADRVLRGSR LGAVSYETDR NHDLAPRQVA RYRTDNGEEF DVPFADDAEI PGTWLCRNGL EGTLIEGDVP EPKKVKPPRT HWDMLLERR SVEELEELLK ERLDLIKAKR RGTGS

UniProtKB: RNA polymerase-binding protein RbpA

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Macromolecule #9: PafC

MacromoleculeName: PafC / type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycolicibacterium smegmatis MC2 155 (bacteria)
Molecular weightTheoretical: 34.044402 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MSQVSTRLVR LLNMVPYFQA NPKVTRAEAA AALGVTGKQL DADLDQLWMC GLPGYSPGDL IDFDFVGDTI EVTFSAGVDH PLRLTSTEA TGILVALRAL VDVPGMVDPE AARSAIAKIE SAVGSQRAVV EGITEDTSAE PGAAATVRTA VRENRALTLE Y YSASRDSL ...String:
MSQVSTRLVR LLNMVPYFQA NPKVTRAEAA AALGVTGKQL DADLDQLWMC GLPGYSPGDL IDFDFVGDTI EVTFSAGVDH PLRLTSTEA TGILVALRAL VDVPGMVDPE AARSAIAKIE SAVGSQRAVV EGITEDTSAE PGAAATVRTA VRENRALTLE Y YSASRDSL ATRTVDPIRV VLVGDNSYLE AWCRSAEAVR LFRFDRIVDA QLLDDPAAPP PPAVAAGPDT SLFDADPSLP SA TLLIGAA AAWMFDYYPL RDITERPDGS CEATMTYASE DWMARFILGF GAEVQVLAPE SLATRVRQAA EAALQAYARC V

UniProtKB: Protein pafC

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Macromolecule #10: Transcriptional regulator-like protein

MacromoleculeName: Transcriptional regulator-like protein / type: protein_or_peptide / ID: 10 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycolicibacterium smegmatis MC2 155 (bacteria)
Molecular weightTheoretical: 36.207555 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: GLSAVSKVER LMNLVIALLS TRTYLPAEKI RTTVAGYADS PSDEAFSRMF ERDKNELRDL GIPLETGRVS KWDSTEGYRI NRDSYALPP IGLTADEAAA VAVATQLWQS PELVTATQNA VLKLRAAGVD VDADGVGVAI ASTATLPGVR GSEEVLQSLL S AIDEGRAV ...String:
GLSAVSKVER LMNLVIALLS TRTYLPAEKI RTTVAGYADS PSDEAFSRMF ERDKNELRDL GIPLETGRVS KWDSTEGYRI NRDSYALPP IGLTADEAAA VAVATQLWQS PELVTATQNA VLKLRAAGVD VDADGVGVAI ASTATLPGVR GSEEVLQSLL S AIDEGRAV QFEHRPSRSA DYTTRTVEPW GVVTHRGRWY LVGHDRDRED TRTFRLSRIS AAARPIGPAG AVQKPQDVNL RD IVRRAVA EQPTGERARI WIAGGRATAL RRQAVTSTPR TIGGRAGEEI TVDIGTWDRL AREIASYGSD AVALEPSSLR DDV VERLRA HAAGGER

UniProtKB: Transcriptional regulator-like protein

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Macromolecule #7: recA-op non-template strand

MacromoleculeName: recA-op non-template strand / type: dna / ID: 7 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Mycolicibacterium smegmatis MC2 155 (bacteria)
Molecular weightTheoretical: 21.006416 KDa
SequenceString: (DG)(DT)(DG)(DG)(DT)(DG)(DA)(DA)(DG)(DA) (DG)(DT)(DT)(DC)(DG)(DA)(DC)(DC)(DG)(DG) (DA)(DC)(DT)(DT)(DG)(DT)(DC)(DG)(DG) (DT)(DG)(DG)(DT)(DC)(DT)(DG)(DC)(DT)(DC) (DT) (DA)(DA)(DC)(DG)(DT)(DC) ...String:
(DG)(DT)(DG)(DG)(DT)(DG)(DA)(DA)(DG)(DA) (DG)(DT)(DT)(DC)(DG)(DA)(DC)(DC)(DG)(DG) (DA)(DC)(DT)(DT)(DG)(DT)(DC)(DG)(DG) (DT)(DG)(DG)(DT)(DC)(DT)(DG)(DC)(DT)(DC) (DT) (DA)(DA)(DC)(DG)(DT)(DC)(DA)(DC) (DG)(DG)(DC)(DC)(DA)(DA)(DC)(DC)(DG)(DA) (DT)(DC) (DG)(DG)(DA)(DA)(DC)(DA)(DC) (DC)

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Macromolecule #8: recA-op template strand

MacromoleculeName: recA-op template strand / type: dna / ID: 8 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Mycolicibacterium smegmatis MC2 155 (bacteria)
Molecular weightTheoretical: 20.912395 KDa
SequenceString: (DG)(DG)(DT)(DG)(DT)(DT)(DC)(DC)(DG)(DA) (DT)(DC)(DG)(DG)(DT)(DA)(DC)(DC)(DG)(DG) (DA)(DC)(DA)(DT)(DG)(DT)(DA)(DA)(DA) (DG)(DA)(DG)(DC)(DA)(DG)(DA)(DC)(DC)(DA) (DC) (DC)(DG)(DA)(DC)(DA)(DA) ...String:
(DG)(DG)(DT)(DG)(DT)(DT)(DC)(DC)(DG)(DA) (DT)(DC)(DG)(DG)(DT)(DA)(DC)(DC)(DG)(DG) (DA)(DC)(DA)(DT)(DG)(DT)(DA)(DA)(DA) (DG)(DA)(DG)(DC)(DA)(DG)(DA)(DC)(DC)(DA) (DC) (DC)(DG)(DA)(DC)(DA)(DA)(DG)(DT) (DC)(DC)(DG)(DG)(DT)(DC)(DG)(DA)(DA)(DC) (DT)(DC) (DT)(DT)(DC)(DA)(DC)(DC)(DA) (DC)

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Macromolecule #11: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 11 / Number of copies: 2 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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Macromolecule #12: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 12 / Number of copies: 1 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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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
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 15 sec.
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 95 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 78.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: cryoSPARC / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: OTHER / Details: cryoSPARC Ab-Initio model
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 470245
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC

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