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- EMDB-24792: M. tuberculosis ribosomal RNA methyltransferase TlyA bound to M. ... -

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Basic information

Entry
Database: EMDB / ID: EMD-24792
TitleM. tuberculosis ribosomal RNA methyltransferase TlyA bound to M. smegmatis 50S ribosomal subunit
Map dataMtb TlyA bound to Msg 50S ribosomal subunit.
Sample
  • Complex: Mtb TlyA bound to Msg 50S ribosomal subunit modified at C2144 with SAM analog NM6.
    • Complex: 50S
      • Protein or peptide: x 32 types
      • RNA: x 2 types
    • Complex: TlyA
      • Protein or peptide: x 1 types
  • Ligand: x 2 types
Function / homology
Function and homology information


23S rRNA (cytidine1920-2'-O)-methyltransferase / methyltransferase activity / large ribosomal subunit / methylation / rRNA binding / ribosome / structural constituent of ribosome / ribonucleoprotein complex / translation / RNA binding / cytoplasm
Similarity search - Function
Haemolysin A /rRNA methyltransferase TlyA / Ribosomal RNA methyltransferase, FtsJ domain / FtsJ-like methyltransferase / Ribosomal protein L9 signature. / Ribosomal protein L9, bacteria/chloroplast / Ribosomal protein L9, C-terminal / Ribosomal protein L9, C-terminal domain / Ribosomal protein L9, C-terminal domain superfamily / Ribosomal protein L36 signature. / Ribosomal protein L9, N-terminal domain superfamily ...Haemolysin A /rRNA methyltransferase TlyA / Ribosomal RNA methyltransferase, FtsJ domain / FtsJ-like methyltransferase / Ribosomal protein L9 signature. / Ribosomal protein L9, bacteria/chloroplast / Ribosomal protein L9, C-terminal / Ribosomal protein L9, C-terminal domain / Ribosomal protein L9, C-terminal domain superfamily / Ribosomal protein L36 signature. / Ribosomal protein L9, N-terminal domain superfamily / Ribosomal protein L9 / Ribosomal protein L9, N-terminal / Ribosomal protein L9, N-terminal domain / Ribosomal protein L36 / Ribosomal protein L36 superfamily / Ribosomal protein L36 / Ribosomal protein L9/RNase H1, N-terminal / Ribosomal protein L34, conserved site / Ribosomal protein L34 signature. / Ribosomal protein L22, bacterial/chloroplast-type / Ribosomal protein L34 / Ribosomal protein L34 / Ribosomal protein L13, bacterial-type / Ribosomal protein L15, bacterial-type / Ribosomal protein L15, conserved site / Ribosomal protein L15 signature. / Ribosomal protein L22/L17, conserved site / S4 RNA-binding domain / RNA-binding S4 domain / RNA-binding S4 domain superfamily / S4 domain / Ribosomal protein L15 / Ribosomal protein L22 signature. / Ribosomal proteins 50S-L15, 50S-L18e, 60S-L27A / Ribosomal protein L22/L17 / Ribosomal protein L22/L17 superfamily / Ribosomal protein L13 / Ribosomal protein L13 / Ribosomal protein L13 superfamily / Ribosomal protein L22p/L17e / Ribosomal protein L18e/L15P / Ribosomal L18e/L15P superfamily / S-adenosyl-L-methionine-dependent methyltransferase superfamily
Similarity search - Domain/homology
16S/23S rRNA (Cytidine-2'-O)-methyltransferase TlyA / : / : / : / : / : / : / : / : / : ...16S/23S rRNA (Cytidine-2'-O)-methyltransferase TlyA / : / : / : / : / : / : / : / : / : / : / : / : / : / : / : / : / : / : / : / : / : / : / : / : / : / 50S ribosomal protein L9 / 50S ribosomal protein L13 / 50S ribosomal protein L15 / Large ribosomal subunit protein bL36 / Uncharacterized protein / 50S ribosomal protein L22 / 50S ribosomal protein L34
Similarity search - Component
Biological speciesMycolicibacterium smegmatis (bacteria) / Mycobacterium tuberculosis (bacteria) / Mycobacterium smegmatis (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.05 Å
AuthorsLaughlin ZT / Dunham CM / Conn GL
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R01-AI088025 United States
Citation
Journal: Proc Natl Acad Sci U S A / Year: 2022
Title: 50S subunit recognition and modification by the ribosomal RNA methyltransferase TlyA.
Authors: Zane T Laughlin / Suparno Nandi / Debayan Dey / Natalia Zelinskaya / Marta A Witek / Pooja Srinivas / Ha An Nguyen / Emily G Kuiper / Lindsay R Comstock / Christine M Dunham / Graeme L Conn /
Abstract: Changes in bacterial ribosomal RNA (rRNA) methylation status can alter the activity of diverse groups of ribosome-targeting antibiotics. These modifications are typically incorporated by a single ...Changes in bacterial ribosomal RNA (rRNA) methylation status can alter the activity of diverse groups of ribosome-targeting antibiotics. These modifications are typically incorporated by a single methyltransferase that acts on one nucleotide target and rRNA methylation directly prevents drug binding, thereby conferring drug resistance. Loss of intrinsic methylation can also result in antibiotic resistance. For example, Mycobacterium tuberculosis becomes sensitized to tuberactinomycin antibiotics, such as capreomycin and viomycin, due to the action of the intrinsic methyltransferase TlyA. TlyA is unique among antibiotic resistance-associated methyltransferases as it has dual 16S and 23S rRNA substrate specificity and can incorporate cytidine-2′-O-methylations within two structurally distinct contexts. Here, we report the structure of a mycobacterial 50S subunit-TlyA complex trapped in a postcatalytic state with a S-adenosyl-L-methionine analog using single-particle cryogenic electron microscopy. Together with complementary functional analyses, this structure reveals critical roles in 23S rRNA substrate recognition for conserved residues across an interaction surface that spans both TlyA domains. These interactions position the TlyA active site over the target nucleotide C2144, which is flipped from 23S Helix 69 in a process stabilized by stacking of TlyA residue Phe157 on the adjacent A2143. Base flipping may thus be a common strategy among rRNA methyltransferase enzymes, even in cases where the target site is accessible without such structural reorganization. Finally, functional studies with 30S subunit suggest that the same TlyA interaction surface is employed to recognize this second substrate, but with distinct dependencies on essential conserved residues.
#1: Journal: Biorxiv / Year: 2022
Title: 50S subunit recognition and modification by the Mycobacterium tuberculosis ribosomal RNA methyltransferase TlyA
Authors: Laughlin ZT / Nandi S / Dey D / Zelinskaya N / Witek MA / Srinivas P / Nguyen HA / Kuiper EG / Comstock LR / Dunham CM / Conn GL
History
DepositionAug 31, 2021-
Header (metadata) releaseMar 30, 2022-
Map releaseMar 30, 2022-
UpdateApr 13, 2022-
Current statusApr 13, 2022Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_24792.png

Downloads & links

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Map

FileDownload / File: emd_24792.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMtb TlyA bound to Msg 50S ribosomal subunit.
Voxel sizeX=Y=Z: 1.069 Å
Density
Contour LevelBy AUTHOR: 0.0179
Minimum - Maximum-0.09661683 - 0.16254294
Average (Standard dev.)0.0008408555 (±0.008895291)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions280280280
Spacing280280280
CellA=B=C: 299.32 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Mtb TlyA bound to H69 and proximal rRNA...

Fileemd_24792_additional_1.map
AnnotationMtb TlyA bound to H69 and proximal rRNA without the rest of Msg 50S. Created from multi body refinement and used in model building and refinement.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Msg 50S alone without H69 or Mtb TlyA....

Fileemd_24792_additional_2.map
AnnotationMsg 50S alone without H69 or Mtb TlyA. Created from multi body refinement.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Mtb TlyA bound to Msg 50S ribosomal subunit modified at C2144 wit...

EntireName: Mtb TlyA bound to Msg 50S ribosomal subunit modified at C2144 with SAM analog NM6.
Components
  • Complex: Mtb TlyA bound to Msg 50S ribosomal subunit modified at C2144 with SAM analog NM6.
    • Complex: 50S
      • Protein or peptide: ribosomal protein bL37
      • RNA: 23S rRNA23S ribosomal RNA
      • Protein or peptide: 50S ribosomal protein L2
      • Protein or peptide: 50S ribosomal protein L3
      • Protein or peptide: 50S ribosomal protein L4
      • Protein or peptide: 50S ribosomal protein L5
      • Protein or peptide: 50S ribosomal protein L6
      • Protein or peptide: 50S ribosomal protein L9
      • Protein or peptide: 50S ribosomal protein L10
      • Protein or peptide: 50S ribosomal protein L11
      • Protein or peptide: 50S ribosomal protein L13
      • Protein or peptide: 50S ribosomal protein L14
      • Protein or peptide: 50S ribosomal protein L15
      • Protein or peptide: 50S ribosomal protein L16
      • Protein or peptide: 50S ribosomal protein L17
      • Protein or peptide: 50S ribosomal protein L18
      • Protein or peptide: 50S ribosomal protein L19
      • Protein or peptide: 50S ribosomal protein L20
      • Protein or peptide: 50S ribosomal protein L21
      • Protein or peptide: 50S ribosomal protein L22
      • Protein or peptide: 50S ribosomal protein L23
      • Protein or peptide: 50S ribosomal protein L24
      • Protein or peptide: 50S ribosomal protein L25
      • Protein or peptide: 50S ribosomal protein L27
      • Protein or peptide: 50S ribosomal protein L28
      • Protein or peptide: 50S ribosomal protein L29
      • Protein or peptide: 50S ribosomal protein L30
      • Protein or peptide: 50S ribosomal protein L32
      • Protein or peptide: 50S ribosomal protein L33
      • Protein or peptide: 50S ribosomal protein L34
      • Protein or peptide: 50S ribosomal protein L35
      • Protein or peptide: 50S ribosomal protein L36
      • Protein or peptide: 50S ribosomal protein L31
      • RNA: 5S rRNA5S ribosomal RNA
    • Complex: TlyA
      • Protein or peptide: 16S/23S rRNA (Cytidine-2'-O)-methyltransferase TlyA
  • Ligand: MAGNESIUM ION
  • Ligand: ZINC ION

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Supramolecule #1: Mtb TlyA bound to Msg 50S ribosomal subunit modified at C2144 wit...

SupramoleculeName: Mtb TlyA bound to Msg 50S ribosomal subunit modified at C2144 with SAM analog NM6.
type: complex / ID: 1 / Parent: 0
Macromolecule list: #1-#2, #6, #8-#14, #16-#20, #22-#25, #27-#29, #31-#32
Details: TlyA occupancy on 50S promoted by covalent modification of NM6 to modification site (C2144).

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Supramolecule #2: 50S

SupramoleculeName: 50S / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1, #3-#35
Source (natural)Organism: Mycolicibacterium smegmatis (bacteria) / Strain: LR222 C101A

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Supramolecule #3: TlyA

SupramoleculeName: TlyA / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2
Source (natural)Organism: Mycobacterium tuberculosis (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)

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Macromolecule #1: ribosomal protein bL37

MacromoleculeName: ribosomal protein bL37 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 2.710179 KDa
SequenceString:
AKRGRKKRDR KHSKANHGKR PNA

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Macromolecule #2: 16S/23S rRNA (Cytidine-2'-O)-methyltransferase TlyA

MacromoleculeName: 16S/23S rRNA (Cytidine-2'-O)-methyltransferase TlyA / type: protein_or_peptide / ID: 2
Details: No density was seen for the His-tag and first three residues (MAR) of TlyA so they were not modeled.
Number of copies: 1 / Enantiomer: LEVO / EC number: 23S rRNA (cytidine1920-2'-O)-methyltransferase
Source (natural)Organism: Mycobacterium tuberculosis (bacteria)
Molecular weightTheoretical: 29.898172 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MHHHHHHASG LVPRGSMARR ARVDAELVRR GLARSRQQAA ELIGAGKVRI DGLPAVKPAT AVSDTTALTV VTDSERAWVS RGAHKLVGA LEAFAIAVAG RRCLDAGAST GGFTEVLLDR GAAHVVAADV GYGQLAWSLR NDPRVVVLER TNARGLTPEA I GGRVDLVV ...String:
MHHHHHHASG LVPRGSMARR ARVDAELVRR GLARSRQQAA ELIGAGKVRI DGLPAVKPAT AVSDTTALTV VTDSERAWVS RGAHKLVGA LEAFAIAVAG RRCLDAGAST GGFTEVLLDR GAAHVVAADV GYGQLAWSLR NDPRVVVLER TNARGLTPEA I GGRVDLVV ADLSFISLAT VLPALVGCAS RDADIVPLVK PQFEVGKGQV GPGGVVHDPQ LRARSVLAVA RRAQELGWHS VG VKASPLP GPSGNVEYFL WLRTQTDRAL SAKGLEDAVH RAISEGP

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Macromolecule #4: 50S ribosomal protein L2

MacromoleculeName: 50S ribosomal protein L2 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 30.008379 KDa
SequenceString: GIRKYKPTTP GRRGASVSDF AEITRSTPEK SLVRPLHGKG GRNAHGRITT RHKGGGHKRA YRVIDFRRHD KDGVNAKVAH IEYDPNRTA NIALLHYLDG EKRYIIAPQG LKQGDVIESG ANADIKPGNN LPLRNIPAGT VIHAVELRPG GGAKLARSAG V SIQLLGKE ...String:
GIRKYKPTTP GRRGASVSDF AEITRSTPEK SLVRPLHGKG GRNAHGRITT RHKGGGHKRA YRVIDFRRHD KDGVNAKVAH IEYDPNRTA NIALLHYLDG EKRYIIAPQG LKQGDVIESG ANADIKPGNN LPLRNIPAGT VIHAVELRPG GGAKLARSAG V SIQLLGKE GTYAALRMPS GEIRRVDVRC RATVGEVGNA EQSNINWGKA GRMRWKGKRP TVRGVVMNPV DHPHGGGEGK TS GGRHPVS PWGKPEGRTR KPNKPSDKLI VRRRRTGK

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Macromolecule #5: 50S ribosomal protein L3

MacromoleculeName: 50S ribosomal protein L3 / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 22.621902 KDa
SequenceString: ARKGILGTKL GMTQVFDENN KVVPVTVVKA GPNVVTRIRT TERDGYSAVQ LAYGEISPRK VIKPVAGQFA AAGVNPRRHV AELRLDDEA AVAEYEVGQE LTAEIFSDGA YVDVTGTSKG KGFAGTMKRH GFRGQGAAHG AQAVHRRPGS IGGCATPGRV F KGTRMSGR ...String:
ARKGILGTKL GMTQVFDENN KVVPVTVVKA GPNVVTRIRT TERDGYSAVQ LAYGEISPRK VIKPVAGQFA AAGVNPRRHV AELRLDDEA AVAEYEVGQE LTAEIFSDGA YVDVTGTSKG KGFAGTMKRH GFRGQGAAHG AQAVHRRPGS IGGCATPGRV F KGTRMSGR MGNDRVTTQN LKVHKVDAEN GVLLIKGAIP GRNGGLVVVR SAIKRG

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Macromolecule #6: 50S ribosomal protein L4

MacromoleculeName: 50S ribosomal protein L4 / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 22.477496 KDa
SequenceString: MTLKVDVKTP AGKTDGSVEL PAELFDVEPN IALMHQVVTA QLAAKRQGTH STKTRGEVSG GGKKPYRQKG TGRARQGSTR APQFTGGGT VHGPKPRDYS QRTPKKMIAA ALRGALSDRA RNDRIHAVTE LVEGQTPSTK SAKTFLGTLT ENKKVLVVIG R TDEVGAKS ...String:
MTLKVDVKTP AGKTDGSVEL PAELFDVEPN IALMHQVVTA QLAAKRQGTH STKTRGEVSG GGKKPYRQKG TGRARQGSTR APQFTGGGT VHGPKPRDYS QRTPKKMIAA ALRGALSDRA RNDRIHAVTE LVEGQTPSTK SAKTFLGTLT ENKKVLVVIG R TDEVGAKS VRNLPGVHVI SPDQLNTYDV LNADDVVFSV EALNAYISAN SK

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Macromolecule #7: 50S ribosomal protein L5

MacromoleculeName: 50S ribosomal protein L5 / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 20.588701 KDa
SequenceString:
KALPRLKQRY REEIREALQQ EFNYANVMQI PGVVKVVVNM GVGDAARDAK LINGAINDLA LITGQKPEVR RARKSIAQFK LREGMPIGA RVTLRGDRMW EFLDRLISIA LPRIRDFRGL SPKQFDGTGN YTFGLNEQSM FHEIDVDSID RPRGMDITVV T TATNDAEG RALLRALGFP FKEN

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Macromolecule #8: 50S ribosomal protein L6

MacromoleculeName: 50S ribosomal protein L6 / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 19.165934 KDa
SequenceString:
SRIGKQPVPV PSGVDVTING QNLSVKGPKG TLTLDVAEPI SVSRAEDGAI VVTRPDDERR SRSLHGLSRT LIANLVTGVT EGYTQKMEI FGVGYRVQLK GQNLEFALGY SHPVLIEAPE GITFAVESPT KFSVSGIDKQ KVGQISAVIR RLRRPDPYKG K GVRYEGEQ IRRKVGKT

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Macromolecule #9: 50S ribosomal protein L9

MacromoleculeName: 50S ribosomal protein L9 / type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 15.949253 KDa
SequenceString:
MKLILTAEVE HLGAAGDTVE VKDGYGRNYL LPRGLAIVAS RGAERQAEEI RRARESKVIR DIEHANELKT ALEGLGDVTL SVNAAGDTG KLFGSVTAAD VVNAIKKAGG PNLDKRTVQL AKAHIKSVGT HPVTVKLHTG VEAKVSLNVV AQ

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Macromolecule #10: 50S ribosomal protein L10

MacromoleculeName: 50S ribosomal protein L10 / type: protein_or_peptide / ID: 10 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 13.095021 KDa
SequenceString:
MAKADKATAV ADIAEQFKAS TATVVTEYRG LTVANLAELR RALGDSATYT VAKNTLVKRA ASEAGIEGLD ELFAGPTAIA FVKGEAVDA AKAIKKFAKD NKALVIKGGY MDGKALSVAD VEKIADL

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Macromolecule #11: 50S ribosomal protein L11

MacromoleculeName: 50S ribosomal protein L11 / type: protein_or_peptide / ID: 11 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 14.109185 KDa
SequenceString:
LIKLQIQAGQ ANPAPPVGPA LGQHGVNIME FCKAYNAATE SQRGNVIPVE ITVYEDRSFT FALKTPPAAK LLLKAAGVQK GSGEPHKTK VAKVTWDQVR EIAETKKADL NANDIDAAAK IIAGTARSMG ITVE

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Macromolecule #12: 50S ribosomal protein L13

MacromoleculeName: 50S ribosomal protein L13 / type: protein_or_peptide / ID: 12 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 16.015493 KDa
SequenceString:
PTYTPKAGDT TRSWYVIDAS DVVLGRLASA AATLLRGKHK PTFTPNVDGG DFVIVINADK IAVSGDKLTK KFAYRHSGYP GGLRKRTIG ELLEKHPTRV VENAIIGMLP HNKLGRQIQK KLKVYAGPDH PHAAQQPIPF EIKQVAQ

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Macromolecule #13: 50S ribosomal protein L14

MacromoleculeName: 50S ribosomal protein L14 / type: protein_or_peptide / ID: 13 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 13.400625 KDa
SequenceString:
MIQQESRLKV ADNTGAKEIL CIRVLGGSSR RYAGIGDVIV ATVKDAIPGG NVKRGDVVKA VVVRTVKERR RADGSYIKFD ENAAVIIKN DNDPRGTRIF GPVGRELREK KFMKIVSLAP EVL

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Macromolecule #14: 50S ribosomal protein L15

MacromoleculeName: 50S ribosomal protein L15 / type: protein_or_peptide / ID: 14 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 15.383968 KDa
SequenceString:
VIKLHDLKPA PGEKKAKTRV GRGEGSKGKT AGRGTKGTKA RKNVPVMFEG GQMPIHMRLP KLKGFKNRFR TEYQVVNVGD INKAFPQGG TVGVDELVAK GLVRKNSLVK VLGDGKLTVK VDVTANKFSG SAREAITAAG GSATEL

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Macromolecule #15: 50S ribosomal protein L16

MacromoleculeName: 50S ribosomal protein L16 / type: protein_or_peptide / ID: 15 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 15.498937 KDa
SequenceString:
MLIPRKVKHR KQHHPEQRGI ASGGTSVSFG DYGIQALEHA YITNRQIESA RIAINRHIKR GGKVWINIFP DRPLTKKPAE TRMGSGKGS PEWWVANVKP GRVLFELSYP DEKTARDALT RAIHKLPIKA RIVTREE

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Macromolecule #16: 50S ribosomal protein L17

MacromoleculeName: 50S ribosomal protein L17 / type: protein_or_peptide / ID: 16 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 13.210346 KDa
SequenceString:
PKPTKGPRLG GSSSHQSALL ANLATSLFEH GRIKTTEPKA RALRPYAEKL ITHAKKGALH NRREVMKKIR DKDVVHTLFA EIGPFYADR NGGYTRIIKV ENRKGDNAPM AVIELVREK

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Macromolecule #17: 50S ribosomal protein L18

MacromoleculeName: 50S ribosomal protein L18 / type: protein_or_peptide / ID: 17 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 13.580531 KDa
SequenceString:
AHKPVGQNIS EVRRNARLRR HARLRKKVAG TAEVPRLVVN RSARHIHVQL VNDLNGTTLA AASSIEADVR AIDGDKKAHS VRVGQLIAE RAKAAGVETV VFDRGGYTYG GRIAALADAA REAGLKF

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Macromolecule #18: 50S ribosomal protein L19

MacromoleculeName: 50S ribosomal protein L19 / type: protein_or_peptide / ID: 18 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 12.892806 KDa
SequenceString:
MNTLDFVDQA SLRDDIPTFS PGDTVNVHVK VIEGSKERIQ VFKGVVIRRQ GGGISETFTV RKESYGVGVE RTFPVHSPNI DHIDVLTRG DVRRAKLYYL RELRGKKAKI KEKR

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Macromolecule #19: 50S ribosomal protein L20

MacromoleculeName: 50S ribosomal protein L20 / type: protein_or_peptide / ID: 19 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 14.035175 KDa
SequenceString:
ARVKRALNAQ KKRRTVLKAS KGYRGQRSRL YRKAKEQQLH SLTYAYRDRR ARKGEFRKLW ISRINAAARA NDITYNRLIQ GLKAAGVEV DRKNLAELAV SDPAAFTALV DVARAALPED VNAPS

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Macromolecule #20: 50S ribosomal protein L21

MacromoleculeName: 50S ribosomal protein L21 / type: protein_or_peptide / ID: 20 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 10.72444 KDa
SequenceString:
TYAIVKTGGK QYKVAAGDVV KVEKLDSEPG ASVSLPVALV VDGANVTSKA DDLAKVAVTA EVLEHTKGPK IRIHKFKNKT GYHKRQGHR QQLTVLKVTG I

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Macromolecule #21: 50S ribosomal protein L22

MacromoleculeName: 50S ribosomal protein L22 / type: protein_or_peptide / ID: 21 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 12.408192 KDa
SequenceString:
EFPSATAKAR YVRVSATKAR RVIDLVRGKS VEEALDILRW APQAASEPVA KVIASAAANA QNNEGLDPST LVVATVYADE GPTAKRIRP RAQGRAFRIR KRTSHITVIV ESRPP

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Macromolecule #22: 50S ribosomal protein L23

MacromoleculeName: 50S ribosomal protein L23 / type: protein_or_peptide / ID: 22 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 10.748448 KDa
SequenceString:
ATITDPRDII LAPVISEKSY GLIEDNVYTF VVHPDSNKTQ IKIAIEKIFD VKVDSVNTAN RQGKRKRTRT GFGKRKSTKR AIVKLAAGS KPIDLFGA

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Macromolecule #23: 50S ribosomal protein L24

MacromoleculeName: 50S ribosomal protein L24 / type: protein_or_peptide / ID: 23
Details: Residues 48-55 are not shown in model. Little to no density seen there.
Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 11.228946 KDa
SequenceString:
MKVHKGDTVL VISGKDKGAK GKVLVAYPDR NKVLVEGVNR IKKHTAVSAN ERGASSGGIV TQEAPIHVSN VMVVDSDGKP TRVGYRIDD ETGKKVRIAK TNGKDI

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Macromolecule #24: 50S ribosomal protein L25

MacromoleculeName: 50S ribosomal protein L25 / type: protein_or_peptide / ID: 24 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 20.331725 KDa
SequenceString: TANIPNKLTA NVRTRTGKGA SRQARRDGKV PAVLYGHGTD PQHLELNARD FAAVLRSHGT NAILTLDIEG TEQLALTKAL DVHPIRRNI QHADLLVVQR GEKVTVEVTV LVEGDATPGT LVTQDANTIE IEAEALSIPE QLTVSVEGVE AGTQITAGQI S LPEGVNLI ...String:
TANIPNKLTA NVRTRTGKGA SRQARRDGKV PAVLYGHGTD PQHLELNARD FAAVLRSHGT NAILTLDIEG TEQLALTKAL DVHPIRRNI QHADLLVVQR GEKVTVEVTV LVEGDATPGT LVTQDANTIE IEAEALSIPE QLTVSVEGVE AGTQITAGQI S LPEGVNLI SDPELLVVNV VEAPSAEALE EEGA

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Macromolecule #25: 50S ribosomal protein L27

MacromoleculeName: 50S ribosomal protein L27 / type: protein_or_peptide / ID: 25 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 8.322479 KDa
SequenceString:
SSSRNGRDSA AQRLGVKRFG GQVVKAGEIL VRQRGTHFHP GVNVGRGGDD TLFALAPGAV EFGAKRGRKT VNIVPVARP

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Macromolecule #26: 50S ribosomal protein L28

MacromoleculeName: 50S ribosomal protein L28 / type: protein_or_peptide / ID: 26 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 6.75982 KDa
SequenceString:
AAVCDICGKG PGFGKSVSHS HRRTSRRWNP NIQPVRAVTR PGGNKQRINA CTSCIKAGKV SRA

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Macromolecule #27: 50S ribosomal protein L29

MacromoleculeName: 50S ribosomal protein L29 / type: protein_or_peptide / ID: 27 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 7.590576 KDa
SequenceString:
GTTPGELREL TDDELKDKLR ESKEELFNLR FQMATGQLSN NRRLRTVRQE IARVYTVLRE RELG

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Macromolecule #28: 50S ribosomal protein L30

MacromoleculeName: 50S ribosomal protein L30 / type: protein_or_peptide / ID: 28 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 6.761772 KDa
SequenceString:
AELKITQVRS TIGARWKQRE SLRTLGLKKI RQSVVREDNA QTRGLINTVH HLVEVEEVG

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Macromolecule #29: 50S ribosomal protein L32

MacromoleculeName: 50S ribosomal protein L32 / type: protein_or_peptide / ID: 29 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 6.050149 KDa
SequenceString:
AVPKRRMSRA NTRSRRAQWK AEAPGLVTVS VAGQQRKVPR RLLKAARLGL VDLD

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Macromolecule #30: 50S ribosomal protein L33

MacromoleculeName: 50S ribosomal protein L33 / type: protein_or_peptide / ID: 30 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 5.833775 KDa
SequenceString:
DVRPKITLAC EVCKHRNYIT KKNRRNDPDR LEIKKFCPNC GTHQPHKES

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Macromolecule #31: 50S ribosomal protein L34

MacromoleculeName: 50S ribosomal protein L34 / type: protein_or_peptide / ID: 31 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 5.391384 KDa
SequenceString:
AKGKRTFQPN NRRRARVHGF RLRMRTRAGR AIVANRRSKG RRALTA

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Macromolecule #32: 50S ribosomal protein L35

MacromoleculeName: 50S ribosomal protein L35 / type: protein_or_peptide / ID: 32 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 7.16731 KDa
SequenceString:
PKAKTHSGAS KRFRRTGTGK IVRQKANRRH LLEHKPTKRT RRLDGRTTVS AADNSRINKL LNG

+
Macromolecule #33: 50S ribosomal protein L36

MacromoleculeName: 50S ribosomal protein L36 / type: protein_or_peptide / ID: 33 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: ATCC 700084 / mc(2)155
Molecular weightTheoretical: 4.341277 KDa
SequenceString:
MKVNPSVKPI CDKCRVIRRH GRVMVICSDP RHKQRQG

+
Macromolecule #34: 50S ribosomal protein L31

MacromoleculeName: 50S ribosomal protein L31 / type: protein_or_peptide / ID: 34 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 5.257952 KDa
SequenceString:
MKTGIHPEYV DTTVQCGCGH SFTTRSTKQS GTIVVEVCSQ CHPFYTGK

+
Macromolecule #3: 23S rRNA

MacromoleculeName: 23S rRNA / type: rna / ID: 3 / Number of copies: 1
Source (natural)Organism: Mycolicibacterium smegmatis (bacteria)
Molecular weightTheoretical: 1.0125324379999998 MDa
SequenceString: UAAGUGUUUA AGGGCGCAUG GUGGAUGCCU UGGCACUGGG AGCCGAUGAA GGACGUAGGA GGCUGCGAUA AGCCUCGGGG AGCUGUCAA CCGAGCGUUG AUCCGAGGAU GUCCGAAUGG GGAAACCCGG CACGAGUGAU GUCGUGUCAC CAGGCGCUGA A UAUAUAGG ...String:
UAAGUGUUUA AGGGCGCAUG GUGGAUGCCU UGGCACUGGG AGCCGAUGAA GGACGUAGGA GGCUGCGAUA AGCCUCGGGG AGCUGUCAA CCGAGCGUUG AUCCGAGGAU GUCCGAAUGG GGAAACCCGG CACGAGUGAU GUCGUGUCAC CAGGCGCUGA A UAUAUAGG CGUCUGGGGG GAACGCGGGG AAGUGAAACA UCUCAGUACC CGUAGGAAGA GAAAACAAAA UGUGAUUCCG UG AGUAGUG GCGAGCGAAA GCGGAGGAUG GCUAAACCGU AUGCAUGUGA UACCGGGUAG GGGUUGUGUG UGCGGGGUUG UGG GACCUA UCUUUCCGGC UCUACCUGGC UGGAGGGCAG UGAGAAAAUG UUGUGGUUAG CGGAAAUGGC UUGGGAUGGC CUGC CGUAG ACGGUGAGAG CCCGGUACGU GAAAACCCGA CGUCUGUCUU GAUGGUGUUC CCGAGUAGCA GCGGGCCCGU GGAAU CUGC UGUGAAUCUG CCGGGACCAC CCGGUAAGCC UGAAUACUUC CCAGUGACCG AUAGCGGAUU AGUACCGUGA GGGAAU GGU GAAAAGUACC CCGGGAGGGG AGUGAAAGAG UACCUGAAAC CGUGCGCUUA CAAUCCGUCA GAGCCCUCGA CGUGUCG UG GGGUGAUGGC GUGCCUUUUG AAGAAUGAGC CUGCGAGUCA GGGACAUGUC GCGAGGUUAA CCCGGGUGGG GUAGCCGC A GCGAAAGCGA GUCUGAAUAG GGCGUAUCCA CACAAGAGUG UGUGGUGUAG UGGUGUGUUC UGGACCCGAA GCGGAGUGA UCUACCCAUG GCCAGGGUGA AGCGCGGGUA AGACCGCGUG GAGGCCCGAA CCCACUUAGG UUGAAGACUG AGGGGAUGAG CUGUGGGUA GGGGUGAAAG GCCAAUCAAA CUCCGUGAUA GCUGGUUCUC CCCGAAAUGC AUUUAGGUGC AGCGUCGCAU G UUUCUUGC CGGAGGUAGA GCUACUGGAU GGCCGAUGGG CCCCACAGGG UUACUGACGU CAGCCAAACU CCGAAUGCCG GU AAGUCCA AGAGUGCGGC AGUGAGACGG CGGGGGAUAA GCUCCGUGCG UCGAGAGGGA AACAGCCCAG AUCGCCGGCU AAG GCCCCU AAGCGUGUGC UAAGUGGAAA AGGAUGUGCA GUCGCGAAGA CAACCAGGAG GUUGGCUUAG AAGCAGCCAC CCUU GAAAG AGUGCGUAAU AGCUCACUGG UCAAGUGAUU GUGCGCCGAU AAUGUAGCGG GGCUCAAGCA CACCGCCGAA GCCGC GGCA GCCAACGUGU UGGCUGGGUA GGGGAGCGUC CUGCAUCCGG UGAAGCCGCC GAGUGAUCGA GUGGUGGAGG GUGUGG GAG UGAGAAUGCA GGCAUGAGUA GCGAUUAGGC AAGUGAGAAC CUUGCCCGCC GAAAGACCAA GGGUUCCUGG GCCAGGC CA GUCCGCCCAG GGUGAGUCGG GACCUAAGGC GAGGCCGACA GGCGUAGUCG AUGGACAACG GGUUGAUAUU CCCGUACC C GUGUAUGUGC GUCCAUGAUG AAUCAGCGGU ACUAACCAUC CAAAACCACC GUGACCGCAC CUUUCGGGGU GUGGCGUUG GUGGGGCUGC AUGGGACCUU CGUUGGUAGU AGUCAAGCGA UGGGGUGACG CAGGAAGGUA GCCGUACCGG UCAGUGGUAA UACCGGGGU AAGCCUGUAG GGAGUCAGAU AGGUAAAUCC GUCUGGCAUA UAUCCUGAGA GGUGAUGCAU AGCCGAGUGA G GCGAAUUC GGUGAUCCUA UGCUGCCGAG AAAAGCCUCU AGCGAGGACA UACACGGCCC GUACCCCAAA CCAACACAGG UG GUCAGGU AGAGAAUACU AAGGCGUACG AGUGAACUAU GGUUAAGGAA CUCGGCAAAA UGCCCCCGUA ACUUCGGGAG AAG GGGGAC CCACAUGGCG UGUAAGCCUU UACGGCCCAA GCGUGAGUGG GUGGCACAAA CCAGUGAGAA GCGACUGUUU ACUA AAAAC ACAGGUCCGU GCGAAGUCGC AAGACGAUGU AUACGGACUG ACGCCUGCCC GGUGCUGGAA GGUUAAGAGG ACCCG UUAA CUCCCUUUGG GGGUGAAGCG GAGAAUUUAA GCCCCAGUAA ACGGCGGUGG UAACUAUAA(AI5) CAUCCUAAGG UA GCGAAAU UCCUUGUCGG GUAAGUUCCG ACCUGCACGA AUGGCGUAAC GACUUCUCAA CUGUCUCAAC CAUAGACUCG GCG AAAUUG CACUACGAGU AAAGAUGCUC GUUACGCGCG GCAGGACGAA AAGACCCCGG GACCUUCACU ACAACUUGGU AUUG GUGCU CGAUACGGUU UGUGUAGGAU AGGUGGGAGA CUGUGAAGCU CACACGCCAG UGUGGGUGGA GUCGUUGUUG AAAUA CCAC UCUGAUCGUA UUGGGCCUCU AACCUCGGAC CGUAUAUCCG GUUCAGGGAC AGUGCCUGGU GGGUAGUUUA ACUGGG GCG GUUGCCUCCU AAAAUGUAAC GGAGGCGCCC AAAGGUUCCC UCAACCUGGA CGGCAAUCAG GUGUUGAGUG UAAGUGC AC AAGGGAGCUU GACUGCGAGA CGGACAUGUC GAGCAGGGAC GAAAGUCGGG ACUAGUGAUC CGGCACCUCU GAGUGGAA G GGGUGUCGCU CAACGGAUAA AAGGUACCCC GGGGAUAACA GGCUGAUCUU CCCCAAGAGU CCAUAUCGAC GGGAUGGUU UGGCACCUCG AUGUCGGCUC GUCGCAUCCU GGGGCUGGAG CAGGUCCCAA GGGUUGGGCU GUUCGCCCAU UAAAGCGGCA CGCGAGCUG GGUUUAGAAC GUCGUGAGAC AGUUCGGUCU CUAUCCGCCG CGCGCGUCAG AAGCUUGAGG AAACCUGUCC C UAGUACGA GAGGACCGGG ACGGACGAAC CUCUGGUAUA CCAGUUGUCC CACCAGGGGC ACGGCUGGAU AGCCACGUUC GG ACAGGAU AACCGCUGAA AGCAUCUAAG CGGGAAACCU CUUCCAAGAC CAGGCUUCUC ACCCUCUAGG AGGGAUAAGG CCC CCCGCA GACCACGGGA UUGAUAGACC AGACCUGGAA GCCUAGUAAU AGGUGCAGGG AACUGGCACU AACCGGCCGA AAAC UUAC

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Macromolecule #35: 5S rRNA

MacromoleculeName: 5S rRNA / type: rna / ID: 35 / Number of copies: 1
Source (natural)Organism: Mycolicibacterium smegmatis (bacteria) / Strain: LR222 C101A
Molecular weightTheoretical: 38.061816 KDa
SequenceString:
GUUACGGCGG UCCAUAGCGG CAGGGAAACG CCCGGUCCCA UCCCGAACCC GGAAGCUAAG CCUGCCAGCG CCGAUGAUAC UACCCAUCC GGGUGGAAAA GUAGGACACC GCCGAACAC

+
Macromolecule #36: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 36 / Number of copies: 402 / Formula: MG
Molecular weightTheoretical: 24.305 Da

+
Macromolecule #37: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 37 / Number of copies: 4 / Formula: ZN
Molecular weightTheoretical: 65.409 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: 7
Component:
ConcentrationName
20.0 mMHEPES
5.0 mMMagnesium Acetate
2.0 mMAmmonium Chloride
75.0 mMPotassium Chloride
3.0 mMBeta-mercaptoethanol2-Mercaptoethanol

Details: Both components dialyzed into this buffer
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 298 K / Instrument: GATAN CRYOPLUNGE 3 / Details: 3.0 or 3.3 second blot time allowed for sample.
Details0.5 micromolar 50S, 5 micromolar TlyA, 10 micromolar NM6

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Electron microscopy

MicroscopeTFS KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 81000
Sample stageCooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 2 / Number real images: 3364 / Average exposure time: 2.0 sec. / Average electron dose: 50.79 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

-
Image processing

Particle selectionNumber selected: 1016454
CTF correctionSoftware - Name: Gctf
Startup modelType of model: EMDB MAP
EMDB ID:

3133

Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final 3D classificationNumber classes: 3 / Avg.num./class: 122800 / Software - Name: RELION (ver. 3.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.05 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 129011
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model
PDB IDChain

5o60


3hp7

chain_id: B

5kyg

chain_id: B
Output model

PDB-7s0s:
M. tuberculosis ribosomal RNA methyltransferase TlyA bound to M. smegmatis 50S ribosomal subunit

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