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BMRB: 18334

2LQX

NMR spatial structure of the trypsin inhibitor BWI-2c from the buckwheat seeds

Authors

Mineev, K.S., Vassilevski, A.A., Oparin, P.B., Grishin, E.V., Egorov, T.A.

Assembly

trypsin inhibitor BWI-2c

Entity

1. trypsin inhibitor BWI-2c (polymer, Thiol state: all disulfide bound), 41 monomers, 5185.850 Da Detail

SEKPQQELEE CQNVCRMKRW STEMVHRCEK KCEEKFERQQ R

Formula weight

5185.85 Da

Entity Connection

disulfide 2 Detail

ID	Type	Value order	Atom ID 1	Atom ID 2
1	disulfide	sing	1:CYS11:SG	1:CYS32:SG
2	disulfide	sing	1:CYS15:SG	1:CYS28:SG

Source organism

Fagopyrum esculentum

Exptl. method

Refine. method

simulated annealing

Data set

assigned_chemical_shifts

Chem. Shift Complete

Sequence coverage: 100.0 %, Completeness: 64.9 %, Completeness (bb): 51.2 % Detail

Polymer type: polypeptide(L)

	Total	¹H	¹³C	¹⁵N
All	64.9 % (338 of 521)	93.7 % (268 of 286)	14.9 % (28 of 188)	89.4 % (42 of 47)
Backbone	51.2 % (125 of 244)	93.8 % (76 of 81)	10.6 % (13 of 123)	90.0 % (36 of 40)
Sidechain	66.0 % (210 of 318)	89.8 % (184 of 205)	18.9 % (20 of 106)	85.7 % (6 of 7)
Aromatic	53.8 % (14 of 26)	100.0 % (13 of 13)	0.0 % (0 of 12)	100.0 % (1 of 1)
Methyl	100.0 % (14 of 14)	100.0 % (7 of 7)	100.0 % (7 of 7)

1. bwi2c

SEKPQQELEE CQNVCRMKRW STEMVHRCEK KCEEKFERQQ R

Show sample condition

Sample #1

Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 273 K, pH 6.8 (±0.1), Details sample in water

#	Name	Isotope labeling	Type	Concentration
1	bwi2c	[U-100% 15N]		1 mM
2	sodium azide	natural abundance		1 mM
3	sodium chloride	natural abundance		20 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

Sample #2

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 6.8 (±0.1), Details sample in d2o

#	Name	Isotope labeling	Type	Concentration
6	bwi2c	natural abundance		1 mM
7	sodium azide	natural abundance		1 mM
8	sodium chloride	natural abundance		20 mM
9	D2O	natural abundance		100 %

Protein Blocks Logo

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

Release date

2012-05-28

Citation

Buckwheat trypsin inhibitor with helical hairpin structure belongs to a new family of plant defence peptides
Oparin, P.B., Mineev, K.S., Dunaevsky, Y.E., Arseniev, A.S., Belozersky, M.A., Grishin, E.V., Egorov, T.A., Vassilevski, A.A.
Biochem. J. (2012), 446, 69-77, PubMed 22612157 , DOI 10.1042/BJ20120548 , Abstract

Entries sharing articles Swiss-Prot: 1 entries Detail

Swiss-Prot: P86794 released on 2012-09-05
Title ITR2C_FAGES Entity Trypsin inhibitor 2c

Related entities 1. trypsin inhibitor BWI-2c, : 1 : 1 : 8 entities Detail

Experiments performed 9 experiments Detail

1 2D 1H-15N HSQC

Instrument

Bruker Avance - 700 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 273 K, pH 6.8 (±0.1), Details sample in water

#	Name	Isotope labeling	Type	Concentration
1	bwi2c	[U-100% 15N]		1 mM
2	sodium azide	natural abundance		1 mM
3	sodium chloride	natural abundance		20 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

2 3D 1H-15N NOESY

Instrument

Bruker Avance - 700 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 273 K, pH 6.8 (±0.1), Details sample in water

#	Name	Isotope labeling	Type	Concentration
1	bwi2c	[U-100% 15N]		1 mM
2	sodium azide	natural abundance		1 mM
3	sodium chloride	natural abundance		20 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

3 3D 1H-15N TOCSY

Instrument

Bruker Avance - 700 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 273 K, pH 6.8 (±0.1), Details sample in water

#	Name	Isotope labeling	Type	Concentration
1	bwi2c	[U-100% 15N]		1 mM
2	sodium azide	natural abundance		1 mM
3	sodium chloride	natural abundance		20 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

4 3D HNHA

Instrument

Bruker Avance - 700 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 273 K, pH 6.8 (±0.1), Details sample in water

#	Name	Isotope labeling	Type	Concentration
1	bwi2c	[U-100% 15N]		1 mM
2	sodium azide	natural abundance		1 mM
3	sodium chloride	natural abundance		20 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

5 3D HNHB

Instrument

Bruker Avance - 700 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 273 K, pH 6.8 (±0.1), Details sample in water

#	Name	Isotope labeling	Type	Concentration
1	bwi2c	[U-100% 15N]		1 mM
2	sodium azide	natural abundance		1 mM
3	sodium chloride	natural abundance		20 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

6 2D DQF-COSY

Instrument

Bruker Avance - 700 MHz

Sample

State isotropic, Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 6.8 (±0.1), Details sample in d2o

#	Name	Isotope labeling	Type	Concentration
6	bwi2c	natural abundance		1 mM
7	sodium azide	natural abundance		1 mM
8	sodium chloride	natural abundance		20 mM
9	D2O	natural abundance		100 %

7 2D 1H-1H TOCSY

Instrument

Bruker Avance - 700 MHz

Sample

State isotropic, Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 6.8 (±0.1), Details sample in d2o

#	Name	Isotope labeling	Type	Concentration
6	bwi2c	natural abundance		1 mM
7	sodium azide	natural abundance		1 mM
8	sodium chloride	natural abundance		20 mM
9	D2O	natural abundance		100 %

8 2D 1H-1H NOESY

Instrument

Bruker Avance - 700 MHz

Sample

State isotropic, Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 6.8 (±0.1), Details sample in d2o

#	Name	Isotope labeling	Type	Concentration
6	bwi2c	natural abundance		1 mM
7	sodium azide	natural abundance		1 mM
8	sodium chloride	natural abundance		20 mM
9	D2O	natural abundance		100 %

9 2D 1H-13C HSQC

Instrument

Bruker Avance - 700 MHz

Sample

State isotropic, Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 6.8 (±0.1), Details sample in d2o

#	Name	Isotope labeling	Type	Concentration
6	bwi2c	natural abundance		1 mM
7	sodium azide	natural abundance		1 mM
8	sodium chloride	natural abundance		20 mM
9	D2O	natural abundance		100 %

NMR combined restraints 8 contents Detail

Properties

Input source #1: NMR data (NEF) - Assigned chemical shifts, Distance restraints, Dihedral angle restraints	combined_18334_2lqx.nef
Input source #2: Coordindates	2lqx.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:11:CYS:SG	A:32:CYS:SG	unknown	unknown	1.974
A:15:CYS:SG	A:28:CYS:SG	unknown	unknown	2.036

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--------40-
SEKPQQELEECQNVCRMKRWSTEMVHRCEKKCEEKFERQQR
|||||||||||||||||||||||||||||||||||||||||
SEKPQQELEECQNVCRMKRWSTEMVHRCEKKCEEKFERQQR

Chain assignments

Entity_assembly_ID (NMR)	Auth_asym_ID (model)	Length	Unmapped	Conflict	Sequence coverage (%)
A	A	41	0	0	100.0

Content subtype: combined_18334_2lqx.nef

#	Content subtype	Saveframe	Status	# of rows (sets)	Experiment type	Sequence coverage (%)
1	Covalent bonds	assembly	OK	2		No information
1	Assigned chemical shifts	assigned_chem_shift_list_1	Warning	334		100.0 (chain: A, length: 41)
1	Distance restraints	DYANA/DIANA_distance_constraints_2	Warning	340 (1)	noe	95.1 (chain: A, length: 41)
2	Distance restraints	DYANA/DIANA_distance_constraints_3	Warning	6 (1)	disulfide_bond	9.8 (chain: A, length: 41)
3	Distance restraints	DYANA/DIANA_distance_constraints_5	Warning	6 (1)	disulfide_bond	9.8 (chain: A, length: 41)
4	Distance restraints	DYANA/DIANA_distance_constraints_4	OK	45 (1)	hbond	56.1 (chain: A, length: 41)
5	Distance restraints	DYANA/DIANA_distance_constraints_6	Warning	45 (1)	hbond	56.1 (chain: A, length: 41)
1	Dihedral angle restraints	DYANA/DIANA_dihedral_8	OK	427 (427)	.	100.0 (chain: A, length: 41)

Assigned chemical shifts

Saveframe: assigned_chem_shift_list_1
- Status: Warning
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 41, Sequence coverage: 100.0%
- Total number of assignments
  - ¹H chemical shifts: 268
  - ¹³C chemical shifts: 21
  - ¹⁵N chemical shifts: 45
- Completeness of assignments
  - Entity_assemble_ID: A
- 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_2
- Status: Warning
- Experiment type: noe
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 41, Sequence coverage: 95.1%
- Total number of restraints: 340
- Weight of restraints: 1.0 (340)
- Potential type of restraints: upper-bound-parabolic (340)
- Range of distance target values: 0.0, 5.5 (Å)
- Histogram of 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: disulfide_bond
  - Sequence coverage of experimental data
    - Entity_assembly_ID: A, Chain length: 41, Sequence coverage: 9.8%
  - Total number of restraints: 6
  - Weight of restraints: 1.0 (6)
  - Potential type of restraints: upper-bound-parabolic (6)
  - Range of distance target values: 2.1, 3.1 (Å)
  - Histogram of 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: disulfide_bond
    - Sequence coverage of experimental data
      - Entity_assembly_ID: A, Chain length: 41, Sequence coverage: 9.8%
    - Total number of restraints: 6
    - Weight of restraints: 1.0 (6)
    - Potential type of restraints: upper-bound-parabolic (6)
    - Range of distance target values: 2.0, 3.0 (Å)
    - Histogram of distance target values
    - Distance restraints per residue
      - Chain_ID: A
    - Distance restraints on contact map
    - Saveframe: DYANA/DIANA_distance_constraints_4
      - Status: OK
      - Experiment type: hbond
      - Sequence coverage of experimental data
        Entity_assembly_ID: A, Chain length: 41, Sequence coverage: 56.1%
        --------10--------20--------30--------40- SEKPQQELEECQNVCRMKRWSTEMVHRCEKKCEEKFERQQR |||||||||| ||||||||||||| ......ELEECQNVCR......EMVHRCEKKCEEK --------10--------20--------30-----
      - Total number of restraints: 45
        Medium range restraints, 1 < | i - j | < 5 : 15
        Backbone-backbone: 15
        Total hydrogen bond restraints: 30
        O...H-x: 15
        O...h-N: 15
      - Weight of restraints: 1.0 (45)
        Medium range restraints, 1 < | i - j | < 5 : 1.0 (15)
        Backbone-backbone: 1.0 (15)
        Total hydrogen bond restraints: 1.0 (30)
        O...H-x: 1.0 (15)
        O...h-N: 1.0 (15)
      - Potential type of restraints: upper-bound-parabolic (45)
        Medium range restraints, 1 < | i - j | < 5 : upper-bound-parabolic (15)
        Backbone-backbone: upper-bound-parabolic (15)
        Total hydrogen bond restraints: upper-bound-parabolic (30)
        O...H-x: upper-bound-parabolic (15)
        O...h-N: upper-bound-parabolic (15)
      - Range of distance target values: 2.2, 3.3 (Å)
      - Histogram of distance target values
      - Distance restraints per residue
        Chain_ID: A
      - Distance restraints on contact map
        Chain_ID_1: A - Chain_ID_2: A
      - Saveframe: DYANA/DIANA_distance_constraints_6
        Status: Warning
        Experiment type: hbond
        Sequence coverage of experimental data
        Entity_assembly_ID: A, Chain length: 41, Sequence coverage: 56.1%
        --------10--------20--------30--------40- SEKPQQELEECQNVCRMKRWSTEMVHRCEKKCEEKFERQQR |||||||||| ||||||||||||| ......ELEECQNVCR......EMVHRCEKKCEEK --------10--------20--------30-----
        Total number of restraints: 45
        Medium range restraints, 1 < | i - j | < 5 : 15
        Backbone-backbone: 15
        Total hydrogen bond restraints: 30
        O...H-x (too close!): 15
        O...h-N (too close!): 15
        Weight of restraints: 1.0 (45)
        Medium range restraints, 1 < | i - j | < 5 : 1.0 (15)
        Backbone-backbone: 1.0 (15)
        Total hydrogen bond restraints: 1.0 (30)
        O...H-x (too close!): 1.0 (15)
        O...h-N (too close!): 1.0 (15)
        Potential type of restraints: upper-bound-parabolic (45)
        Medium range restraints, 1 < | i - j | < 5 : upper-bound-parabolic (15)
        Backbone-backbone: upper-bound-parabolic (15)
        Total hydrogen bond restraints: upper-bound-parabolic (30)
        O...H-x (too close!): upper-bound-parabolic (15)
        O...h-N (too close!): upper-bound-parabolic (15)
        Range of distance target values: 1.7, 2.6 (Å)
        Histogram of distance target values
        Distance restraints per residue
        Chain_ID: A
        Distance restraints on contact map
        Chain_ID_1: A - Chain_ID_2: A

Dihedral angle restraints

Saveframe: DYANA/DIANA_dihedral_8
- Status: OK
- Experiment type: .
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 41, Sequence coverage: 100.0%
- Total number of restraints: 427
- Total number of restraints per polymer type: 427
- Weight of restraints: 1.0 (427)
- Potential type of restraints: square-well-parabolic (427)
- Plot of Phi-Psi angle restraints
- Plot of Chi1-Chi2 angle restraints
- Dihedral angle restraints per residue
  - Chain_ID: A

Keywords buckwheat, BWI-2c, helical hairpin, NMR, trypsin inhibitor