PDB-1np9: Structure of the parallel-stranded DNA quadruplex d(TTAGGGA)4 con...

Basic information

• Entry: Database: PDB / ID: 1np9
• Title: Structure of the parallel-stranded DNA quadruplex d(TTAGGGA)4 containing the human telomeric repeat
• Components: 5'-D(*TP*TP*AP*GP*GP*GP*T)-3'
• Keywords: DNA / PARALLEL-STRANDED QUADRUPLEX DNA / TTAGGGT REPEAT / A-TETRAD
• Function / homology: DNA
• Method: SOLUTION NMR / restrained molecular dynamics
• Model type details: minimized average
• Authors: Gavathiotis, E. / Searle, M.S.
• Citation: Journal: ORG.BIOMOL.CHEM. / Year: 2003; Title: Structure of the parallel-stranded DNA quadruplex d(TTAGGGT)4 containing the human telomeric repeat: evidence for A-tetrad formation from NMR and molecular dynamics simulations.; Authors: Gavathiotis, E. / Searle, M.S.

History

Deposition	Jan 17, 2003	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Sep 16, 2003	Provider: repository / Type: Initial release
Revision 1.1	Apr 29, 2008	Group: Version format compliance
Revision 1.2	Jul 13, 2011	Group: Version format compliance
Revision 1.3	Feb 23, 2022	Group: Data collection / Database references / Derived calculations Category: database_2 / pdbx_nmr_software / pdbx_struct_assembly / pdbx_struct_oper_list Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_nmr_software.name
Revision 1.4	May 22, 2024	Group: Data collection / Category: chem_comp_atom / chem_comp_bond

Assembly

Deposited unit

A: 5'-D(*TP*TP*AP*GP*GP*GP*T)-3'

B: 5'-D(*TP*TP*AP*GP*GP*GP*T)-3'

C: 5'-D(*TP*TP*AP*GP*GP*GP*T)-3'

D: 5'-D(*TP*TP*AP*GP*GP*GP*T)-3'

Summary:

• 8.67 kDa, 4 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	8,674	4
Polymers	8,674	4
Non-polymers	0	0
Water	0	0

1

Summary:

• Idetical with deposited unit
• defined by author

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1

NMR ensembles

	Data	Criteria
Number of conformers (submitted / calculated)	11 / 100	structures with the lowest energy
Representative	Model #11	minimized average structure

Components

#1: DNA chain

A B C D
5'-D(*TP*TP*AP*GP*GP*GP*T)-3'

Details:

Mass: 2168.445 Da / Num. of mol.: 4 / Source method: obtained synthetically

Experimental details

Experiment

• Experiment: Method: SOLUTION NMR

NMR experiment

Conditions-ID	Experiment-ID	Solution-ID	Type
1	1	1	2D NOESY
1	2	1	2D TOCSY
1	3	1	DQF-COSY

• NMR details: Text: Standard phase sensitive 2D NMR pulse sequences were used.

Sample preparation

• Details: Contents: 6.4mM TTAGGGT, 100mM KCl, 10mM K2HPO4, 0.1 mM EDTA, 0.1 mM NaN3, 90% H2O, 10% D2O; Solvent system: 90% H2O/10% D2O
• Sample conditions: pH: 7.0 / Pressure: 1 atm / Temperature: 298 K
• Crystal grow: Method: other / Details: NMR

NMR measurement

• Radiation: Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M
• Radiation wavelength: Relative weight: 1

NMR spectrometer

Type	Manufacturer	Model	Field strength (MHz)	Spectrometer-ID
Bruker DRX	Bruker	DRX	500	1
Bruker DRX	Bruker	DRX	600	2

Processing

NMR software

Name	Version	Classification
XwinNMR	2.6	collection
XwinNMR	2.6	processing
ANSIG	v3.3	data analysis
Amber	6	refinement

• Refinement: Method: restrained molecular dynamics / Software ordinal: 1 ; Details: Structures calculated using restrained molecular dynamics.Total number of NOE restraints 728 of which 24 hydrogen-bond restraints included for the hydrogen-bonding geometry of the G-tetrads. Energy minimisations and restrained molecular dynamics were carried out using the SANDER module of AMBER 6. Calculations with SANDER were performed with a 2 fs time step, with the SHAKE algorithm (tolerance 0.00005 A) applied to all bonds to remove bond stretching, and a 9 A cut off to the Lennard Jones interactions. The restrained molecular dynamics were performed at 300K and a constant pressure of 1.0 atm with isotropic position scaling utilising the Berendsen algorithm for temperature coupling. Translational and rotational motions were removed every 100 fs. All calculations were carried out with the PME method using a 9 A cut-off for direct space non-bonded calculations and a 0.00001 Ewald convergence tolerance for the inclusion of long-range electrostatics in our calculations. The quadruplex system was allowed to equilibrate fully before the molecular dynamics calculations. Minimisation was performed with 50 steps of steepest descent and 5000 steps of conjugate gradient to first the water and counterions, with the DNA coordinates frozen, followed by a further 5000 steps on all the components of the system. Next, 10 ps unrestrained molecular dynamics were run at 100K on the water alone with the DNA and potassium ions constrained, followed for another 10 ps to allow the potassium ions to move. In the following 5 ps of dynamics the temperature of the system was increased from 100K to 300K. The next runs, each of them of 10 ps dynamics, the DNA force constant is gradually reduced from 100 to 50, 25, 10, 5 and 2.5 kcal mol-1 A-2. The equilibration step ends with 100 ps of dynamics on the whole fully unrestrained system. The system now is fully equilibrated and NOE restraints can be applied to the quadruplex system. Distance restraints were introduced gradually on the system over the first 10 ps of 100 ps MD run with the temperature stable at 300K and PME on. All NOE restraints were introduced in the form of square well potentials with a force constant of 50 kcal mol-1 A-1 for the hydrogen-bond restraints and 30 kcal mol-1 A-1 for all the other NOE distance restraints. A total of 1000 ps simulation was performed under the same conditions.Calculated structures satisfied the vast majority of the NOE restraints from the set of 728 restraints. The average minimised structure had no restraint violation > 0.3 A that contributed to a 48.22 kcal mol-1 energy penalty. Snapshots of each picosecond were extracted from the whole simulation and the structures were determined to be equilibrated on the basis of RMSD analysis.
• NMR representative: Selection criteria: minimized average structure
• NMR ensemble: Conformer selection criteria: structures with the lowest energy; Conformers calculated total number: 100 / Conformers submitted total number: 11