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BMRB: 30011

5I2P

Resonance assignments and NMR structure determination of tarantula toxin- W7A mutant of mu-TRTX-Pre1a

Authors

Chin, Y., Wingerd, J.S., Mobli, M., Rash, L.D.

Assembly

W7A mu-TRTX-Pre1a Toxin

Entity

1. W7A mu-TRTX-Pre1a Toxin (polymer, Thiol state: all disulfide bound), 36 monomers, 4207.832 Da Detail

SEDCLGAFSR CSPKNDKCCP NYKCSSKDLW CKYKIW

Formula weight

4207.832 Da

Entity Connection

disulfide 3 Detail

ID	Type	Value order	Atom ID 1	Atom ID 2
1	disulfide	sing	1:CYS4:SG	1:CYS19:SG
2	disulfide	sing	1:CYS11:SG	1:CYS24:SG
3	disulfide	sing	1:CYS18:SG	1:CYS31:SG

Source organism

Exptl. method

Refine. method

torsion angle dynamics

Data set

assigned_chemical_shifts, spectral_peak_list

Chem. Shift Complete

Sequence coverage: 100.0 %, Completeness: 95.6 %, Completeness (bb): 94.3 % Detail

Polymer type: polypeptide(L)

	Total	¹H	¹³C	¹⁵N
All	95.6 % (412 of 431)	97.8 % (224 of 229)	92.7 % (152 of 164)	94.7 % (36 of 38)
Backbone	94.3 % (200 of 212)	97.2 % (69 of 71)	91.6 % (98 of 107)	97.1 % (33 of 34)
Sidechain	96.9 % (246 of 254)	98.1 % (155 of 158)	95.7 % (88 of 92)	75.0 % (3 of 4)
Aromatic	100.0 % (50 of 50)	100.0 % (25 of 25)	100.0 % (23 of 23)	100.0 % (2 of 2)
Methyl	100.0 % (14 of 14)	100.0 % (7 of 7)	100.0 % (7 of 7)

1. entity 1

SEDCLGAFSR CSPKNDKCCP NYKCSSKDLW CKYKIW

Show sample condition

Sample

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 298 K, pH 6, Details 300 uM [U-99% 13C; U-99% 15N] W7A-mu-TRTX-Pre1a, 10 mM sodium phosphate, 5 % D2O, 95% H2O/5% D2O

#	Name	Isotope labeling	Type	Concentration
1	D2O	natural abundance		5 %
2	W7A-mu-TRTX-Pre1a	[U-99% 13C; U-99% 15N]		300 uM
3	sodium phosphate	natural abundance		10 mM

Protein Blocks Logo

Calculated from 20 models in PDB: 5I2P, Strand ID: A Detail

Release date

2016-02-29

Citation

Resonance assignments and NMR structure determination of tarantula toxin- W7A mutant of mu-TRTX-Pre1a
Wingerd, J.S., Chin, Y., Cristofori-Armstrong, B., Mobli, M., Rash, L.D.

Related entities 1. W7A mu-TRTX-Pre1a Toxin, : 1 : 1 : 97 entities Detail

Experiments performed 9 experiments Detail

1 2D 1H-15N HSQC

Instrument

Bruker AvanceII - 900 MHz nmr900

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 298 K, pH 6, Details 300 uM [U-99% 13C; U-99% 15N] W7A-mu-TRTX-Pre1a, 10 mM sodium phosphate, 5 % D2O, 95% H2O/5% D2O

#	Name	Isotope labeling	Type	Concentration
1	D2O	natural abundance		5 %
2	W7A-mu-TRTX-Pre1a	[U-99% 13C; U-99% 15N]		300 uM
3	sodium phosphate	natural abundance		10 mM

2 3D CBCA(CO)NH

Instrument

Bruker AvanceII - 900 MHz nmr900

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 298 K, pH 6, Details 300 uM [U-99% 13C; U-99% 15N] W7A-mu-TRTX-Pre1a, 10 mM sodium phosphate, 5 % D2O, 95% H2O/5% D2O

#	Name	Isotope labeling	Type	Concentration
1	D2O	natural abundance		5 %
2	W7A-mu-TRTX-Pre1a	[U-99% 13C; U-99% 15N]		300 uM
3	sodium phosphate	natural abundance		10 mM

3 3D HNCO

Instrument

Bruker AvanceII - 900 MHz nmr900

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 298 K, pH 6, Details 300 uM [U-99% 13C; U-99% 15N] W7A-mu-TRTX-Pre1a, 10 mM sodium phosphate, 5 % D2O, 95% H2O/5% D2O

#	Name	Isotope labeling	Type	Concentration
1	D2O	natural abundance		5 %
2	W7A-mu-TRTX-Pre1a	[U-99% 13C; U-99% 15N]		300 uM
3	sodium phosphate	natural abundance		10 mM

4 3D HNCACB

Instrument

Bruker AvanceII - 900 MHz nmr900

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 298 K, pH 6, Details 300 uM [U-99% 13C; U-99% 15N] W7A-mu-TRTX-Pre1a, 10 mM sodium phosphate, 5 % D2O, 95% H2O/5% D2O

#	Name	Isotope labeling	Type	Concentration
1	D2O	natural abundance		5 %
2	W7A-mu-TRTX-Pre1a	[U-99% 13C; U-99% 15N]		300 uM
3	sodium phosphate	natural abundance		10 mM

5 3D 1H-15N NOESY

Instrument

Bruker AvanceII - 900 MHz nmr900

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 298 K, pH 6, Details 300 uM [U-99% 13C; U-99% 15N] W7A-mu-TRTX-Pre1a, 10 mM sodium phosphate, 5 % D2O, 95% H2O/5% D2O

#	Name	Isotope labeling	Type	Concentration
1	D2O	natural abundance		5 %
2	W7A-mu-TRTX-Pre1a	[U-99% 13C; U-99% 15N]		300 uM
3	sodium phosphate	natural abundance		10 mM

6 4D HCC(co)NH-TOCSY

Instrument

Bruker AvanceII - 900 MHz nmr900

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 298 K, pH 6, Details 300 uM [U-99% 13C; U-99% 15N] W7A-mu-TRTX-Pre1a, 10 mM sodium phosphate, 5 % D2O, 95% H2O/5% D2O

#	Name	Isotope labeling	Type	Concentration
1	D2O	natural abundance		5 %
2	W7A-mu-TRTX-Pre1a	[U-99% 13C; U-99% 15N]		300 uM
3	sodium phosphate	natural abundance		10 mM

7 3D 1H-13C NOESY aromatic

Instrument

Bruker AvanceII - 900 MHz nmr900

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 298 K, pH 6, Details 300 uM [U-99% 13C; U-99% 15N] W7A-mu-TRTX-Pre1a, 10 mM sodium phosphate, 5 % D2O, 95% H2O/5% D2O

#	Name	Isotope labeling	Type	Concentration
1	D2O	natural abundance		5 %
2	W7A-mu-TRTX-Pre1a	[U-99% 13C; U-99% 15N]		300 uM
3	sodium phosphate	natural abundance		10 mM

8 3D 1H-13C NOESY aliphatic

Instrument

Bruker AvanceII - 900 MHz nmr900

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 298 K, pH 6, Details 300 uM [U-99% 13C; U-99% 15N] W7A-mu-TRTX-Pre1a, 10 mM sodium phosphate, 5 % D2O, 95% H2O/5% D2O

#	Name	Isotope labeling	Type	Concentration
1	D2O	natural abundance		5 %
2	W7A-mu-TRTX-Pre1a	[U-99% 13C; U-99% 15N]		300 uM
3	sodium phosphate	natural abundance		10 mM

9 3D HBHA(CO)NH

Instrument

Bruker AvanceII - 900 MHz nmr900

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 298 K, pH 6, Details 300 uM [U-99% 13C; U-99% 15N] W7A-mu-TRTX-Pre1a, 10 mM sodium phosphate, 5 % D2O, 95% H2O/5% D2O

#	Name	Isotope labeling	Type	Concentration
1	D2O	natural abundance		5 %
2	W7A-mu-TRTX-Pre1a	[U-99% 13C; U-99% 15N]		300 uM
3	sodium phosphate	natural abundance		10 mM

NMR combined restraints 6 contents Detail

Properties

Input source #1: NMR data (NEF) - Assigned chemical shifts, Distance restraints, Dihedral angle restraints	combined_30011_5i2p.nef
Input source #2: Coordindates	5i2p.cif
Diamagnetism of the molecular assembly	True (excluding Oxygen atoms)
Whether the assembly has a disulfide bond	True (see coordinates for details)
Whether the assembly has a other bond	None
Whether the assembly contains a cyclic polymer	None
Overall data processing status	Warning

Disulfide bonds

Ptnr_site_1	Ptnr_site_2	Redox_state_prediction_1	Redox_state_prediction_2	Distance (Å)
A:4:CYS:SG	A:19:CYS:SG	oxidized, CA 54.192, CB 43.523 ppm	oxidized, CA 53.628, CB 38.785 ppm	1.959
A:11:CYS:SG	A:24:CYS:SG	oxidized, CA 53.841, CB 45.401 ppm	oxidized, CA 56.666, CB 38.452 ppm	2.102
A:18:CYS:SG	A:31:CYS:SG	oxidized, CA 57.02, CB 39.292 ppm	oxidized, CA 55.291, CB 38.969 ppm	2.1

Other bonds (neither disulfide, covalent nor hydrogen bonds, e.g. Zinc–sulphur bond)

None

Non-standard residues

None

Sequence alignments

Entity_assembly_ID (NMR): A vs Auth_asym_ID (model): A

--------10--------20--------30------
SEDCLGAFSRCSPKNDKCCPNYKCSSKDLWCKYKIW
||||||||||||||||||||||||||||||||||||
SEDCLGAFSRCSPKNDKCCPNYKCSSKDLWCKYKIW

Chain assignments

Entity_assembly_ID (NMR)	Auth_asym_ID (model)	Length	Unmapped	Conflict	Sequence coverage (%)
A	A	36	0	0	100.0

Content subtype: combined_30011_5i2p.nef

#	Content subtype	Saveframe	Status	# of rows (sets)	Experiment type	Sequence coverage (%)
1	Covalent bonds	assembly	OK	3		No information
1	Assigned chemical shifts	assigned_chemical_shifts_1	Warning	416		100.0 (chain: A, length: 36)
1	Distance restraints	DYANA/DIANA_distance_constraints_3	OK	676 (1)	noe	100.0 (chain: A, length: 36)
2	Distance restraints	DYANA/DIANA_distance_constraints_4	Warning	18 (1)	undefined	16.7 (chain: A, length: 36)
1	Dihedral angle restraints	DYANA/DIANA_dihedral_2	OK	348 (348)	.	100.0 (chain: A, length: 36)
2	Dihedral angle restraints	DYANA/DIANA_dihedral_5	OK	60 (60)	.	100.0 (chain: A, length: 36)

Assigned chemical shifts

Saveframe: assigned_chemical_shifts_1
- Status: Warning
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 36, Sequence coverage: 100.0%
- Total number of assignments
  - ¹H chemical shifts: 225
  - ¹³C chemical shifts: 155
  - ¹⁵N chemical shifts: 36
- Completeness of assignments
  - Entity_assemble_ID: A
- Redox state of CYS
- Peptide bond of PRO
- Rotameric state of ILE , LEU, and VAL
- Histogram of normalized assigned chemical shifts
- Random coil index and derived parameters (S² and NMR RMSD)
  - Chain_ID: A

Covalent bonds

Saveframe: assembly
- Status: OK

Distance restraints

Saveframe: DYANA/DIANA_distance_constraints_3
- Status: OK
- Experiment type: noe
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 36, Sequence coverage: 100.0%
- Total number of restraints: 676
- Weight of restraints: 1.0 (676)
- Potential type of restraints: upper-bound-parabolic (676)
- Range of distance target values: 2.2, 5.5 (Å)
- Histogram of distance target values
- Histogram of discrepancy in distance target values
- Distance restraints per residue
  - Chain_ID: A
- Distance restraints on contact map
- Saveframe: DYANA/DIANA_distance_constraints_4
  - Status: Warning
  - Experiment type: undefined
  - Sequence coverage of experimental data
    - Entity_assembly_ID: A, Chain length: 36, Sequence coverage: 16.7%
  - Total number of restraints: 18
  - Weight of restraints: 1.0 (18)
  - Potential type of restraints: upper-bound-parabolic (18)
  - Range of distance target values: 1.96, 3.5 (Å)
  - Histogram of distance target values
  - Histogram of discrepancy in distance target values
  - Distance restraints per residue
    - Chain_ID: A
  - Distance restraints on contact map

Dihedral angle restraints

Saveframe: DYANA/DIANA_dihedral_2
- Status: OK
- Experiment type: .
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 36, Sequence coverage: 100.0%
- Total number of restraints: 348
- Total number of restraints per polymer type: 348
- Weight of restraints: 1.0 (348)
- Potential type of restraints: square-well-parabolic (348)
- Plot of Phi-Psi angle restraints
- Plot of Chi1-Chi2 angle restraints
- Dihedral angle restraints per residue
  - Chain_ID: A
- Saveframe: DYANA/DIANA_dihedral_5
  - Status: OK
  - Experiment type: .
  - Sequence coverage of experimental data
    - Entity_assembly_ID: A, Chain length: 36, Sequence coverage: 100.0%
  - Total number of restraints: 60
  - Total number of restraints per polymer type: 60
  - Weight of restraints: 1.0 (60)
  - Potential type of restraints: square-well-parabolic (60)
  - Plot of Phi-Psi angle restraints
  - Dihedral angle restraints per residue
    - Chain_ID: A

Keywords Spider toxin; Voltage-gated sodium channel modulator; Inhibitor cystein knot; disulfide-rich