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- PDB-6mce: Solution structure of HIV-1 TAR with Tat RNA Binding Domain -

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Basic information

Entry
Database: PDB / ID: 6mce
TitleSolution structure of HIV-1 TAR with Tat RNA Binding Domain
Components
  • Protein Tat
  • TAR RNA
KeywordsTRANSCRIPTION / RNA BINDING PROTEIN
Function / homology
Function and homology information


trans-activation response element binding / Interactions of Tat with host cellular proteins / protein serine/threonine phosphatase inhibitor activity / positive regulation of viral transcription / modulation by virus of host chromatin organization / symbiont-mediated suppression of host translation initiation / evasion of host immune response / molecular sequestering activity / host cell nucleolus / actinin binding ...trans-activation response element binding / Interactions of Tat with host cellular proteins / protein serine/threonine phosphatase inhibitor activity / positive regulation of viral transcription / modulation by virus of host chromatin organization / symbiont-mediated suppression of host translation initiation / evasion of host immune response / molecular sequestering activity / host cell nucleolus / actinin binding / negative regulation of peptidyl-threonine phosphorylation / Pausing and recovery of Tat-mediated HIV elongation / Tat-mediated HIV elongation arrest and recovery / RNA-binding transcription regulator activity / Tat-mediated elongation of the HIV-1 transcript / Formation of HIV-1 elongation complex containing HIV-1 Tat / cyclin binding / positive regulation of transcription elongation by RNA polymerase II / PKR-mediated signaling / host cell cytoplasm / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / protein domain specific binding / DNA-templated transcription / host cell nucleus / extracellular region / metal ion binding
Similarity search - Function
Tat domain superfamily / Immunodeficiency virus transactivating regulatory protein (Tat) / Transactivating regulatory protein (Tat)
Similarity search - Domain/homology
RNA / RNA (> 10) / Protein Tat
Similarity search - Component
Biological speciesHuman immunodeficiency virus 1
Human immunodeficiency virus type 1 group M subtype B
MethodSOLUTION NMR / simulated annealing
AuthorsPham, V.V. / D'Souza, V.M.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM103297-01 United States
Howard Hughes Medical Institute (HHMI)55108516 United States
CitationJournal: Nat Commun / Year: 2018
Title: HIV-1 Tat interactions with cellular 7SK and viral TAR RNAs identifies dual structural mimicry.
Authors: Pham, V.V. / Salguero, C. / Khan, S.N. / Meagher, J.L. / Brown, W.C. / Humbert, N. / de Rocquigny, H. / Smith, J.L. / D'Souza, V.M.
History
DepositionAug 31, 2018Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 31, 2018Provider: repository / Type: Initial release
Revision 1.1Nov 20, 2019Group: Author supporting evidence / Data collection / Category: pdbx_audit_support / pdbx_nmr_software
Item: _pdbx_audit_support.funding_organization / _pdbx_nmr_software.name
Revision 1.2May 1, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: TAR RNA
B: Protein Tat


Theoretical massNumber of molelcules
Total (without water)11,8152
Polymers11,8152
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: isothermal titration calorimetry, Obtain a 1:1 binding ratio in the ITC for the peptide and the RNA under the NMR conditions solved
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area2050 Å2
ΔGint-11 kcal/mol
Surface area6660 Å2
NMR ensembles
DataCriteria
Number of conformers (submitted / calculated)10 / 20structures with acceptable covalent geometry
RepresentativeModel #1fewest violations

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Components

#1: RNA chain TAR RNA


Mass: 9652.762 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Human immunodeficiency virus 1
#2: Protein/peptide Protein Tat / Transactivating regulatory protein


Mass: 2162.533 Da / Num. of mol.: 1 / Fragment: RNA Binding Domain residues 44-60 / Source method: obtained synthetically
Source: (synth.) Human immunodeficiency virus type 1 group M subtype B
References: UniProt: P04608

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Experimental details

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Experiment

ExperimentMethod: SOLUTION NMR
NMR experiment
Conditions-IDExperiment-IDSolution-IDSample stateSpectrometer-IDType
111isotropic12D 1H-1H NOESY
123isotropic12D 1H-1H NOESY
132isotropic12D 1H-15N HSQC
144isotropic12D 1H-13C HSQC
154anisotropic12D 1H-13C HSQC

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Sample preparation

Details
TypeSolution-IDContentsLabelSolvent system
solution10.5 mM TAR RNA with deuterated NTPs in a combo of either AC, GU, or GA, 0.5 mM Tat RNA Binding Domain, 90% H2O/10% D2OTAR:Tat-190% H2O/10% D2O
solution30.5 mM TAR RNA deuterated NTPs in a combo of either AC, GU, or GA, 0.5 mM Tat RNA Binding Domain, 100% D2OTAR:Tat-3100% D2O
solution20.5 mM [U-13C; U-15N] TAR RNA, 0.5 mM Tat RNA Binding Domain, 90% H2O/10% D2OTAR:Tat-290% H2O/10% D2O
solution40.5 mM TAR RNA, 0.5 mM [U-13C; U-15N] Tat RNA Binding Domain selectively labeled in each of the peptides: 1) R49, K50, R57, A58 2) K51, R52, Q54,R55 3) R53, Q54, R56, 100% D2OTAR:Tat-4100% D2O
Sample
Conc. (mg/ml)ComponentIsotopic labelingSolution-ID
0.5 mMTAR RNAnatural abundance1
0.5 mMTat RNA Binding Domainnatural abundance1
0.5 mMTAR RNAnatural abundance3
0.5 mMTat RNA Binding Domainnatural abundance3
0.5 mMTAR RNA[U-13C; U-15N]2
0.5 mMTat RNA Binding Domainnatural abundance2
0.5 mMTAR RNAnatural abundance4
0.5 mMTat RNA Binding Domain[U-13C; U-15N]4
Sample conditionsIonic strength: 10mM NaCl, 70mM NaCl, 0.1mM EDTA mM / Label: TAR Buffer / pH: 5.4 / Pressure: 100000 Pa / Temperature: 298 K

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NMR measurement

NMR spectrometerType: Bruker DMX / Manufacturer: Bruker / Model: DMX / Field strength: 700 MHz

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Processing

NMR software
NameDeveloperClassification
TopSpinBruker Biospincollection
NMRViewJohnson, One Moon Scientificpeak picking
NMRViewJohnson, One Moon Scientificchemical shift assignment
CYANAGuntert, Mumenthaler and Wuthrichdata analysis
CYANAGuntert, Mumenthaler and Wuthrichprocessing
CYANAGuntert, Mumenthaler and Wuthrichstructure calculation
AmberCase, Darden, Cheatham III, Simmerling, Wang, Duke, Luo, and Kollmanrefinement
RefinementMethod: simulated annealing / Software ordinal: 7
NMR representativeSelection criteria: fewest violations
NMR ensembleConformer selection criteria: structures with acceptable covalent geometry
Conformers calculated total number: 20 / Conformers submitted total number: 10

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