2022-11-23
REL
RCSB
RCSB
RCSB
2020-05-06
2020-11-18
2020-11-18
2022-11-23
Cryo-EM structure of Bacillus subtilis RNA Polymerase elongation complex
Newing T
Tolun G
Oakley AJ
Newing TP
Oakley AJ
Miller M
Dawson CJ
Brown SHJ
Bouwer JC
Tolun G
Lewis PJ
Molecular basis for RNA polymerase-dependent transcription complex recycling by the helicase-like motor protein HelD.
Nat Commun
UK
11
6420
6420
2020
33339820
doi:10.1038/s41467-020-20157-5
2041-1723
EMD-21920
associated EM volume
Cryo-EM structure of Bacillus subtilis RNA Polymerase elongation complex
6wvj
FULLOVERLAP
DNA-directed RNA polymerase elongation complex
DNA-directed RNA polymerase elongation complex
0
1
2
3
4
5
6
7
0.344200
DNA-directed RNA polymerase subunit alpha, DNA-directed RNA polymerase subunit beta, DNA-directed RNA polymerase subunit beta', DNA-directed RNA polymerase subunit omega/DNA Complex
1
1
2
3
4
Bacillus subtilis
Escherichia coli BL21(DE3)
pNG1256
DNA, RNA
1
5
6
7
Escherichia coli BL21(DE3)
DNA-directed RNA polymerase subunit alpha
Bacillus subtilis (strain 168)
168
0.034842387
2
Escherichia coli BL21(DE3)
LEVO
MIEIEKPKIETVEISDDAKFGKFVVEPLERGYGTTLGNSLRRILLSSLPGAAVTSIQIDGVLHEFSTIEGVVEDVTTIIL
HIKKLALKIYSDEEKTLEIDVQGEGTVTAADITHDSDVEILNPDLHIATLGENASFRVRLTAQRGRGYTPADANKRDDQP
IGVIPIDSIYTPVSRVSYQVENTRVGQVANYDKLTLDVWTDGSTGPKEAIALGSKILTEHLNIFVGLTDEAQHAEIMVEK
EEDQKEKVLEMTIEELDLSVRSYNCLKRAGINTVQELANKTEEDMMKVRNLGRKSLEEVKAKLEELGLGLRKDD
2.7.7.6
DNA-directed RNA polymerase subunit beta
Bacillus subtilis
0.133847938
1
Escherichia coli BL21(DE3)
LEVO
MTGQLVQYGRHRQRRSYARISEVLELPNLIEIQTSSYQWFLDEGLREMFQDISPIEDFTGNLSLEFIDYSLGEPKYPVEE
SKERDVTYSAPLRVKVRLINKETGEVKDQDVFMGDFPIMTDTGTFIINGAERVIVSQLVRSPSVYFSGKVDKNGKKGFTA
TVIPNRGAWLEYETDAKDVVYVRIDRTRKLPVTVLLRALGFGSDQEILDLIGENEYLRNTLDKDNTENSDKALLEIYERL
RPGEPPTVENAKSLLDSRFFDPKRYDLANVGRYKINKKLHIKNRLFNQRLAETLVDPETGEILAEKGQILDRRTLDKVLP
YLENGIGFRKLYPNGGVVEDEVTLQSIKIFAPTDQEGEQVINVIGNAYIEEEIKNITPADIISSISYFFNLLHGVGDTDD
IDHLGNRRLRSVGELLQNQFRIGLSRMERVVRERMSIQDTNTITPQQLINIRPVIASIKEFFGSSQLSQFMDQTNPLAEL
THKRRLSALGPGGLTRERAGMEVRDVHYSHYGRMCPIETPEGPNIGLINSLSSYAKVNRFGFIETPYRRVDPETGKVTGR
IDYLTADEEDNYVVAQANARLDDEGAFIDDSIVARFRGENTVVSRNRVDYMDVSPKQVVSAATACIPFLENDDSNRALMG
ANMQRQAVPLMQPEAPFVGTGMEYVSGKDSGAAVICKHPGIVERVEAKNVWVRRYEEVDGQKVKGNLDKYSLLKFVRSNQ
GTCYNQRPIVSVGDEVVKGEILADGPSMELGELALGRNVMVGFMTWDGYNYEDAIIMSERLVKDDVYTSIHIEEYESEAR
DTKLGPEEITRDIPNVGEDALRNLDDRGIIRIGAEVKDGDLLVGKVTPKGVTELTAEERLLHAIFGEKAREVRDTSLRVP
HGGGGIIHDVKVFNREDGDELPPGVNQLVRVYIVQKRKISEGDKMAGRHGNKGVISKILPEEDMPYLPDGTPIDIMLNPL
GVPSRMNIGQVLELHMGMAARYLGIHIASPVFDGAREEDVWETLEEAGMSRDAKTVLYDGRTGEPFDNRVSVGIMYMIKL
AHMVDDKLHARSTGPYSLVTQQPLGGKAQFGGQRFGEMEVWALEAYGAAYTLQEILTVKSDDVVGRVKTYEAIVKGDNVP
EPGVPESFKVLIKELQSLGMDVKILSGDEEEIEMRDLEDEEDAKQADGLALSGDEEPEETASADVERDVVTKE
2.7.7.6
DNA-directed RNA polymerase subunit beta'
Bacillus subtilis
0.134444953
1
Escherichia coli BL21(DE3)
LEVO
MLDVNNFEYMNIGLASPDKIRSWSFGEVKKPETINYRTLKPEKDGLFCERIFGPTKDWECHCGKYKRVRYKGVVCDRCGV
EVTRAKVRRERMGHIELAAPVSHIWYFKGIPSRMGLVLDMSPRALEEVIYFASYVVTDPANTPLEKKQLLSEKEYRAYLD
KYGNKFQASMGAEAIHKLLQDIDLVKEVDMLKEELKTSQGQRRTRAIKRLEVLEAFRNSGNKPSWMILDVLPVIPPELRP
MVQLDGGRFATSDLNDLYRRVINRNNRLKRLLDLGAPSIIVQNEKRMLQEAVDALIDNGRRGRPVTGPGNRPLKSLSHML
KGKQGRFRQNLLGKRVDYSGRSVIVVGPHLKMYQCGLPKEMALELFKPFVMKELVEKGLAHNIKSAKRKIERVQPEVWDV
LESVIKEHPVLLNRAPTLHRLGIQAFEPTLVEGRAIRLHPLVCTAYNADFDGDQMAVHVPLSAEAQAEARILMLAAQNIL
NPKDGKPVVTPSQDMVLGNYYLTLERAGAVGEGMVFKNTDEALLAYQNGYVHLHTRVAVAANSLKNVTFTEEQRSKLLIT
TVGKLVFNEILPESFPYMNEPTKSNIEEKTPDRFFLEKGADVKAVIAQQPINAPFKKGILGKIIAEIFKRFHITETSKML
DRMKNLGFKYSTKAGITVGVSDIVVLDDKQEILEEAQSKVDNVMKQFRRGLITEEERYERVISIWSAAKDVIQGKLMKSL
DELNPIYMMSDSGARGNASNFTQLAGMRGLMANPAGRIIELPIKSSFREGLTVLEYFISTHGARKGLADTALKTADSGYL
TRRLVDVAQDVIIRETDCGTDRGILAKPLKEGTETIERLEERLIGRFARKQVKHPETGEVLVNENELIDEDKALEIVEAG
IEEVWIRSAFTCNTPHGVCKRCYGRNLATGSDVEVGEAVGIIAAQSIGEPGTQLTMRTFHTGGVAGDDITQGLPRIQELF
EARNPKGQATITEIDGTVVEINEVRDKQQEIVVQGAVETRSYTAPYNSRLKVAEGDKITRGQVLTEGSIDPKELLKVTDL
TTVQEYLLHEVQKVYRMQGVEIGDKHVEVMVRQMLRKVRVIDAGDTDVLPGTLLDIHQFTEANKKVLLEGNRPATGRPVL
LGITKASLETDSFLSAASFQETTRVLTDAAIKGKRDELLGLKENVIIGKLVPAGTGMMKYRKVKPVSNVQPTDDMVPVE
2.7.7.6
DNA-directed RNA polymerase subunit omega
Bacillus subtilis
0.0077669209999999995
1
Escherichia coli BL21(DE3)
LEVO
MLDPSIDSLMNKLDSKYTLVTVSARRAREMQIKKDQMIEHTISHKYVGKALEEIDAGLLSFEKEDRE
2.7.7.6
DNA (5'-D(*TP*GP*TP*CP*GP*GP*GP*CP*GP*TP*CP*CP*GP*CP*GP*CP*GP*CP*C)-3')
Escherichia coli BL21(DE3)
BL21(DE3)
0.005814720999999999
1
(DT)(DG)(DT)(DC)(DG)(DG)(DG)(DC)(DG)(DT)(DC)(DC)(DG)(DC)(DG)(DC)(DG)(DC)(DC)
DNA
RNA (5'-R(P*GP*GP*CP*GP*CP*GP*CP*G)-3')
Escherichia coli BL21(DE3)
BL21(DE3)
0.0025966170000000003
1
GGCGCGCG
DNA (5'-D(P*AP*CP*GP*CP*CP*CP*GP*AP*CP*A)-3')
Escherichia coli BL21(DE3)
BL21(DE3)
0.0029989839999999997
1
(DA)(DC)(DG)(DC)(DC)(DC)(DG)(DA)(DC)(DA)
DNA
ZINC ION
6.5409e-05
2
ZN
MAGNESIUM ION
2.4305e-05
1
MG
singleParticle
particle
2.50
7.8
20.0
NH2C(CH2OH)3
Tris
150.0
NaCl
Sodium Chloride
10.0
MgCl2
Magnesium Chloride
1.0
(HO2CCH2)2NCH2CH2N(CH2CO2H)2
EDTA
5.0
HSCH2CH(OH)CH(OH)CH2SH
DTT
Quantifoil R1.2/1.3
GOLD
300
GOLD
HOLEY
50.0
GLOW DISCHARGE
AIR
ETHANE
100
298
FEI VITROBOT MARK IV
Sample loading volume ranged between 2 and 3 microlitres. Samples were blotted for 5 seconds prior to vitrification..
FEI TITAN KRIOS
FLOOD BEAM
BRIGHT FIELD
FIELD EMISSION GUN
300
2.7
0.6
2.8000000000000003
59500.0
FEI TITAN KRIOS AUTOGRID HOLDER
NITROGEN
GIF Quantum LS
GATAN K2 SUMMIT (4k x 4k)
COUNTING
3838
3710
1-40
2
5185
9.0
52.2
1
463519
Gctf
The initial model was generated using the "3D initial model" algorithm in RELION 3.1
C1
3.36
FSC 0.143 CUT-OFF
RELION
3.1
58854
MAXIMUM LIKELIHOOD
RELION
3.1
MAXIMUM LIKELIHOOD
RELION
3.1
REAL