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BMRB: 19205

2M7T

Solution NMR Structure of Engineered Cystine Knot Protein 2.5D

Authors

Cochran, F., Das, R.

Assembly

Engineered Cystine Knot Protein 2.5D

Entity

1. Engineered Cystine Knot Protein 2.5D (polymer, Thiol state: all disulfide bound), 33 monomers, 3249.551 Da Detail

GCPQGRGDWA PTSCSQDSDC LAGCVCGPNG FCG

Formula weight

3249.551 Da

Entity Connection

disulfide 3 Detail

ID	Type	Value order	Atom ID 1	Atom ID 2
1	disulfide	sing	1:CYS2:SG	1:CYS24:SG
2	disulfide	sing	1:CYS14:SG	1:CYS26:SG
3	disulfide	sing	1:CYS20:SG	1:CYS32:SG

Exptl. method

Refine. method

molecular dynamics

Data set

assigned_chemical_shifts

Chem. Shift Complete

Sequence coverage: 100.0 %, Completeness: 97.8 %, Completeness (bb): 96.9 % Detail

Polymer type: polypeptide(L)

	Total	¹H	¹³C	¹⁵N
All	97.8 % (307 of 314)	98.8 % (160 of 162)	96.6 % (114 of 118)	97.1 % (33 of 34)
Backbone	96.9 % (186 of 192)	98.6 % (69 of 70)	95.7 % (88 of 92)	96.7 % (29 of 30)
Sidechain	99.3 % (147 of 148)	98.9 % (91 of 92)	100.0 % (52 of 52)	100.0 % (4 of 4)
Aromatic	100.0 % (22 of 22)	100.0 % (11 of 11)	100.0 % (10 of 10)	100.0 % (1 of 1)
Methyl	100.0 % (14 of 14)	100.0 % (7 of 7)	100.0 % (7 of 7)

1. entity

GCPQGRGDWA PTSCSQDSDC LAGCVCGPNG FCG

Show sample condition

Sample

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 288 K, pH 6

#	Name	Isotope labeling	Type	Concentration
1	2.5D	[U-13C; U-15N]		300 uM
2	sodium phosphate	natural abundance		50 mM

Protein Blocks Logo

Calculated from 20 models in PDB: 2M7T, Strand ID: A Detail

Release date

2014-05-04

Citation

Challenging the state of the art in protein structure prediction: Highlights of experimental target structures for the 10th Critical Assessment of Techniques for Protein Structure Prediction Experiment CASP10
Kryshtafovych, A., Moult, J., Bales, P., Bazan, J., Biasini, M., Burgin, A., Chen, C., Cochran, F.V., Craig, T.K., Das, R., Fass, D., Garcia-Doval, C., Herzberg, O., Lorimer, D., Luecke, H., Ma, X., Nelson, D.C., van Raaij, M.J., Rohwer, F., Segall, A., Seguritan, V., Zeth, K., Schwede, T.
Proteins (2014), 82 Suppl 2, 26-42, PubMed 24318984 , DOI 10.1002/prot.24489 , Abstract

Related entities 1. Engineered Cystine Knot Protein 2.5D, : 1 : 1 : 18 entities Detail

Experiments performed 11 experiments Detail

1 2D 1H-15N HSQC

Instrument

Varian VNMRS - 800 MHz

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 288 K, pH 6

#	Name	Isotope labeling	Type	Concentration
1	2.5D	[U-13C; U-15N]		300 uM
2	sodium phosphate	natural abundance		50 mM

2 2D 1H-13C HSQC aliphatic

Instrument

Varian VNMRS - 800 MHz

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 288 K, pH 6

#	Name	Isotope labeling	Type	Concentration
1	2.5D	[U-13C; U-15N]		300 uM
2	sodium phosphate	natural abundance		50 mM

3 3D C(CO)NH

Instrument

Varian VNMRS - 800 MHz

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 288 K, pH 6

#	Name	Isotope labeling	Type	Concentration
1	2.5D	[U-13C; U-15N]		300 uM
2	sodium phosphate	natural abundance		50 mM

4 3D HNCO

Instrument

Varian VNMRS - 800 MHz

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 288 K, pH 6

#	Name	Isotope labeling	Type	Concentration
1	2.5D	[U-13C; U-15N]		300 uM
2	sodium phosphate	natural abundance		50 mM

5 3D HNCACB

Instrument

Varian VNMRS - 800 MHz

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 288 K, pH 6

#	Name	Isotope labeling	Type	Concentration
1	2.5D	[U-13C; U-15N]		300 uM
2	sodium phosphate	natural abundance		50 mM

6 3D H(CCO)NH

Instrument

Varian VNMRS - 800 MHz

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 288 K, pH 6

#	Name	Isotope labeling	Type	Concentration
1	2.5D	[U-13C; U-15N]		300 uM
2	sodium phosphate	natural abundance		50 mM

7 3D 1H-15N NOESY

Instrument

Varian VNMRS - 800 MHz

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 288 K, pH 6

#	Name	Isotope labeling	Type	Concentration
1	2.5D	[U-13C; U-15N]		300 uM
2	sodium phosphate	natural abundance		50 mM

8 3D 1H-13C NOESY aliphatic

Instrument

Varian VNMRS - 800 MHz

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 288 K, pH 6

#	Name	Isotope labeling	Type	Concentration
1	2.5D	[U-13C; U-15N]		300 uM
2	sodium phosphate	natural abundance		50 mM

9 3D 1H-13C NOESY aromatic

Instrument

Varian VNMRS - 800 MHz

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 288 K, pH 6

#	Name	Isotope labeling	Type	Concentration
1	2.5D	[U-13C; U-15N]		300 uM
2	sodium phosphate	natural abundance		50 mM

10 ghnco_LRA

Instrument

Varian VNS - 600 MHz

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 288 K, pH 6

#	Name	Isotope labeling	Type	Concentration
1	2.5D	[U-13C; U-15N]		300 uM
2	sodium phosphate	natural abundance		50 mM

11 2D 1H-13C HSQC aromatic

Instrument

Varian VNMRS - 800 MHz

Sample

State isotropic, Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 288 K, pH 6

#	Name	Isotope labeling	Type	Concentration
1	2.5D	[U-13C; U-15N]		300 uM
2	sodium phosphate	natural abundance		50 mM

NMR combined restraints 5 contents Detail

Properties

Input source #1: NMR data (NEF) - Assigned chemical shifts, Distance restraints	combined_19205_2m7t.nef
Input source #2: Coordindates	2m7t.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:2:CYS:SG	A:24:CYS:SG	oxidized, CA 53.145, CB 41.921 ppm	oxidized, CA 54.902, CB 46.168 ppm	2.017
A:14:CYS:SG	A:26:CYS:SG	oxidized, CA 53.806, CB 47.605 ppm	oxidized, CA 55.199, CB 38.26 ppm	2.029
A:20:CYS:SG	A:32:CYS:SG	oxidized, CA 51.981, CB 39.974 ppm	oxidized, CA 55.43, CB 42.087 ppm	2.022

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---
GCPQGRGDWAPTSCSQDSDCLAGCVCGPNGFCG
|||||||||||||||||||||||||||||||||
GCPQGRGDWAPTSCSQDSDCLAGCVCGPNGFCG

Chain assignments

Entity_assembly_ID (NMR)	Auth_asym_ID (model)	Length	Unmapped	Conflict	Sequence coverage (%)
A	A	33	0	0	100.0

Content subtype: combined_19205_2m7t.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_chem_shift_list_1	OK	307		100.0 (chain: A, length: 33)
1	Distance restraints	DYANA/DIANA_distance_constraints_4	OK	597 (1)	noe	97.0 (chain: A, length: 33)
2	Distance restraints	DYANA/DIANA_distance_constraints_5	Warning	18 (1)	undefined	18.2 (chain: A, length: 33)
3	Distance restraints	DYANA/DIANA_distance_constraints_3	Warning	12 (1)	hbond	18.2 (chain: A, length: 33)

Assigned chemical shifts

Saveframe: assigned_chem_shift_list_1
- Status: OK
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 33, Sequence coverage: 100.0%
- Total number of assignments
  - ¹H chemical shifts: 160
  - ¹³C chemical shifts: 114
  - ¹⁵N chemical shifts: 33
- 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_4
- Status: OK
- Experiment type: noe
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 33, Sequence coverage: 97.0%
- Total number of restraints: 594
- Weight of restraints: 1.0 (594)
- Potential type of restraints: upper-bound-parabolic (594)
- Range of distance target values: 2.58, 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_5
  - Status: Warning
  - Experiment type: undefined
  - Sequence coverage of experimental data
    - Entity_assembly_ID: A, Chain length: 33, Sequence coverage: 18.2%
  - Total number of restraints: 18
  - Weight of restraints: 1.0 (18)
  - Potential type of restraints: upper-bound-parabolic (18)
  - Range of distance target values: 2.0, 3.1 (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_3
    - Status: Warning
    - Experiment type: hbond
    - Sequence coverage of experimental data
      - Entity_assembly_ID: A, Chain length: 33, Sequence coverage: 18.2%
    - Total number of restraints: 12
    - Weight of restraints: 1.0 (12)
    - Potential type of restraints: upper-bound-parabolic (12)
    - Range of distance target values: 1.8, 3.0 (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

Keywords CASP, Cystine Knot, Knottin, NMR, Protein Engineering