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- PDB-4n5s: Ternary complex structure of Klenow fragment of Taq DNA polymeras... -

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Basic information

Entry
Database: PDB / ID: 4n5s
TitleTernary complex structure of Klenow fragment of Taq DNA polymerase I707L mutant (Cs3C KlenTaq) with DNA and ddCTP
Components
  • 5'-D(*AP*A*AP*GP*GP*GP*CP*GP*CP*CP*GP*TP*GP*GP*TP*C)-3'
  • 5'-D(*GP*AP*CP*CP*AP*CP*GP*GP*CP*GP*CP*(DOC))-3'
  • DNA polymerase I, thermostable
KeywordsTRANSFERASE/DNA / DNA polymerase / TRANSFERASE-DNA complex
Function / homology
Function and homology information


nucleoside binding / 5'-3' exonuclease activity / DNA-templated DNA replication / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / DNA repair / DNA binding
Similarity search - Function
Taq polymerase, thermostable, exonuclease region / Taq polymerase, exonuclease / DNA polymerase I-like, H3TH domain / 5'-3' exonuclease, C-terminal SAM fold / 5'-3' exonuclease, alpha-helical arch, N-terminal / 5'-3' exonuclease, N-terminal resolvase-like domain / 5'-3' exonuclease / 5'-3' exonuclease / Taq DNA Polymerase; Chain T, domain 4 / Taq DNA Polymerase; Chain T, domain 4 ...Taq polymerase, thermostable, exonuclease region / Taq polymerase, exonuclease / DNA polymerase I-like, H3TH domain / 5'-3' exonuclease, C-terminal SAM fold / 5'-3' exonuclease, alpha-helical arch, N-terminal / 5'-3' exonuclease, N-terminal resolvase-like domain / 5'-3' exonuclease / 5'-3' exonuclease / Taq DNA Polymerase; Chain T, domain 4 / Taq DNA Polymerase; Chain T, domain 4 / DNA polymerase 1 / Alpha-Beta Plaits - #370 / Helix-hairpin-helix motif, class 2 / Helix-hairpin-helix class 2 (Pol1 family) motifs / 5'-3' exonuclease, C-terminal domain superfamily / DNA polymerase A / DNA polymerase family A / DNA-directed DNA polymerase, family A, conserved site / DNA polymerase family A signature. / DNA-directed DNA polymerase, family A, palm domain / DNA polymerase A domain / PIN-like domain superfamily / Helix-hairpin-helix DNA-binding motif, class 1 / Helix-hairpin-helix DNA-binding motif class 1 / Ribonuclease H-like superfamily/Ribonuclease H / Nucleotidyltransferase; domain 5 / Ribonuclease H superfamily / Ribonuclease H-like superfamily / Alpha-Beta Plaits / DNA/RNA polymerase superfamily / Up-down Bundle / 2-Layer Sandwich / Mainly Alpha / Alpha Beta
Similarity search - Domain/homology
ACETATE ION / 2',3'-DIDEOXYCYTIDINE 5'-TRIPHOSPHATE / : / DNA / DNA (> 10) / DNA polymerase I, thermostable
Similarity search - Component
Biological speciesThermus aquaticus (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.67 Å
AuthorsWu, E.Y.
CitationJournal: Biochemistry / Year: 2015
Title: A Conservative Isoleucine to Leucine Mutation Causes Major Rearrangements and Cold Sensitivity in KlenTaq1 DNA Polymerase.
Authors: Wu, E.Y. / Walsh, A.R. / Materne, E.C. / Hiltner, E.P. / Zielinski, B. / Miller, B.R. / Mawby, L. / Modeste, E. / Parish, C.A. / Barnes, W.M. / Kermekchiev, M.B.
History
DepositionOct 10, 2013Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 21, 2015Provider: repository / Type: Initial release
Revision 1.1Feb 11, 2015Group: Database references
Revision 1.2Sep 20, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / pdbx_struct_conn_angle / struct_conn / struct_ref_seq_dif / struct_site
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_struct_conn_angle.ptnr1_auth_asym_id / _pdbx_struct_conn_angle.ptnr1_auth_comp_id / _pdbx_struct_conn_angle.ptnr1_auth_seq_id / _pdbx_struct_conn_angle.ptnr1_label_asym_id / _pdbx_struct_conn_angle.ptnr1_label_atom_id / _pdbx_struct_conn_angle.ptnr1_label_comp_id / _pdbx_struct_conn_angle.ptnr1_label_seq_id / _pdbx_struct_conn_angle.ptnr2_auth_asym_id / _pdbx_struct_conn_angle.ptnr2_auth_comp_id / _pdbx_struct_conn_angle.ptnr2_auth_seq_id / _pdbx_struct_conn_angle.ptnr2_label_asym_id / _pdbx_struct_conn_angle.ptnr2_label_atom_id / _pdbx_struct_conn_angle.ptnr2_label_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_asym_id / _pdbx_struct_conn_angle.ptnr3_auth_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_seq_id / _pdbx_struct_conn_angle.ptnr3_label_asym_id / _pdbx_struct_conn_angle.ptnr3_label_atom_id / _pdbx_struct_conn_angle.ptnr3_label_comp_id / _pdbx_struct_conn_angle.ptnr3_label_seq_id / _pdbx_struct_conn_angle.value / _struct_conn.pdbx_dist_value / _struct_conn.pdbx_leaving_atom_flag / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id / _struct_ref_seq_dif.details / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
B: 5'-D(*GP*AP*CP*CP*AP*CP*GP*GP*CP*GP*CP*(DOC))-3'
C: 5'-D(*AP*A*AP*GP*GP*GP*CP*GP*CP*CP*GP*TP*GP*GP*TP*C)-3'
A: DNA polymerase I, thermostable
hetero molecules


Theoretical massNumber of molelcules
Total (without water)72,27015
Polymers71,0163
Non-polymers1,25412
Water9,620534
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area8050 Å2
ΔGint-45 kcal/mol
Surface area24910 Å2
MethodPISA
Unit cell
Length a, b, c (Å)107.657, 107.657, 89.659
Angle α, β, γ (deg.)90.00, 90.00, 120.00
Int Tables number152
Space group name H-MP3121

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Components

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DNA chain , 2 types, 2 molecules BC

#1: DNA chain 5'-D(*GP*AP*CP*CP*AP*CP*GP*GP*CP*GP*CP*(DOC))-3'


Mass: 3617.371 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: primer strand
#2: DNA chain 5'-D(*AP*A*AP*GP*GP*GP*CP*GP*CP*CP*GP*TP*GP*GP*TP*C)-3'


Mass: 4964.216 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: template strand

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Protein , 1 types, 1 molecules A

#3: Protein DNA polymerase I, thermostable / Taq polymerase 1


Mass: 62434.773 Da / Num. of mol.: 1 / Fragment: Klenow fragment (UNP residues 281-832) / Mutation: I707L
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Thermus aquaticus (bacteria) / Gene: polA, pol1 / Production host: Escherichia coli (E. coli) / References: UniProt: P19821, DNA-directed DNA polymerase

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Non-polymers , 7 types, 546 molecules

#4: Chemical
ChemComp-MN / MANGANESE (II) ION


Mass: 54.938 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: Mn
#5: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg
#6: Chemical ChemComp-DCT / 2',3'-DIDEOXYCYTIDINE 5'-TRIPHOSPHATE


Type: DNA linking / Mass: 451.158 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C9H16N3O12P3
#7: Chemical ChemComp-EPE / 4-(2-HYDROXYETHYL)-1-PIPERAZINE ETHANESULFONIC ACID / HEPES


Mass: 238.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C8H18N2O4S / Comment: pH buffer*YM
#8: Chemical ChemComp-ACT / ACETATE ION


Mass: 59.044 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C2H3O2
#9: Chemical ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL


Mass: 92.094 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C3H8O3
#10: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 534 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.11 Å3/Da / Density % sol: 41.76 %
Crystal growTemperature: 289 K / Method: vapor diffusion, hanging drop / pH: 7.5
Details: 0.1 M HEPES, pH 7.5, 20 mM manganese chloride, 0.1 M sodium acetate, 10% w/v PEG4000, VAPOR DIFFUSION, HANGING DROP, temperature 289K

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 22-BM / Wavelength: 0.97911 Å
DetectorType: MARMOSAIC 225 mm CCD / Detector: CCD / Date: Nov 18, 2012
RadiationMonochromator: Rosenbaum-Rock double-crystal Si(220) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97911 Å / Relative weight: 1
ReflectionResolution: 1.67→32.8 Å / Num. all: 67931 / Num. obs: 64469 / Observed criterion σ(I): -3 / Redundancy: 3.6 % / Rsym value: 0.047 / Net I/σ(I): 30.19
Reflection shellResolution: 1.67→1.7 Å / Redundancy: 2.7 % / Num. unique all: 2624 / Rsym value: 0.376 / % possible all: 76.3

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Processing

Software
NameVersionClassification
SERGUIdata collection
PHASERphasing
REFMAC5.7.0029refinement
HKL-2000data reduction
HKL-2000data scaling
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 3KTQ

3ktq


Resolution: 1.67→32.8 Å / Cor.coef. Fo:Fc: 0.969 / Cor.coef. Fo:Fc free: 0.96 / SU B: 1.787 / SU ML: 0.061 / Cross valid method: THROUGHOUT / ESU R: 0.098 / ESU R Free: 0.095 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflectionSelection details
Rfree0.19156 3432 5.1 %RANDOM
Rwork0.16104 ---
all0.16259 67931 --
obs0.16259 64469 97.33 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK
Displacement parametersBiso mean: 24.379 Å2
Baniso -1Baniso -2Baniso -3
1--0 Å2-0 Å2-0 Å2
2---0 Å2-0 Å2
3---0 Å2
Refinement stepCycle: LAST / Resolution: 1.67→32.8 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms4286 511 63 534 5394
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0110.0185003
X-RAY DIFFRACTIONr_bond_other_d0.0010.024609
X-RAY DIFFRACTIONr_angle_refined_deg1.5571.8986884
X-RAY DIFFRACTIONr_angle_other_deg0.877310586
X-RAY DIFFRACTIONr_dihedral_angle_1_deg5.5445541
X-RAY DIFFRACTIONr_dihedral_angle_2_deg29.56622.315203
X-RAY DIFFRACTIONr_dihedral_angle_3_deg13.43915757
X-RAY DIFFRACTIONr_dihedral_angle_4_deg19.6041553
X-RAY DIFFRACTIONr_chiral_restr0.1060.2735
X-RAY DIFFRACTIONr_gen_planes_refined0.0090.0215241
X-RAY DIFFRACTIONr_gen_planes_other0.0010.021145
LS refinement shellResolution: 1.67→1.714 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0.254 201 -
Rwork0.229 3775 -
obs-2624 77.88 %

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