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BMRB: 25753

2N66

NMR Structure of sweeter mutant (D40K) of sweet protein Brazzein

Authors

Singarapu, K.K., Tonelli, M., Markley, J.L., Assadi-Porter, F.

Assembly

sweeter mutant (D40K) of sweet protein Brazzein

Entity

1. sweeter mutant (D40K) of sweet protein Brazzein (polymer), 54 monomers, 6511.359 Da Detail

QDKCKKVYEN YPVSKCQLAN QCNYDCKLDK HARSGECFYK EKRNLQCICD YCEY

Formula weight

6511.359 Da

Source organism

Pentadiplandra brazzeana

Exptl. method

Refine. method

distance geometry

Data set

assigned_chemical_shifts

Chem. Shift Complete

Sequence coverage: 96.3 %, Completeness: 81.6 %, Completeness (bb): 80.1 % Detail

Polymer type: polypeptide(L)

	Total	¹H	¹³C	¹⁵N
All	81.6 % (537 of 658)	92.0 % (323 of 351)	63.8 % (157 of 246)	93.4 % (57 of 61)
Backbone	80.1 % (258 of 322)	96.3 % (104 of 108)	64.6 % (104 of 161)	94.3 % (50 of 53)
Sidechain	84.8 % (330 of 389)	90.1 % (219 of 243)	75.4 % (104 of 138)	87.5 % (7 of 8)
Aromatic	45.2 % (28 of 62)	90.3 % (28 of 31)	0.0 % (0 of 31)
Methyl	100.0 % (28 of 28)	100.0 % (14 of 14)	100.0 % (14 of 14)

1. entity

QDKCKKVYEN YPVSKCQLAN QCNYDCKLDK HARSGECFYK EKRNLQCICD YCEY

Show sample condition

Sample

Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 310 K, pH 5.2

#	Name	Isotope labeling	Type	Concentration
1	D40K	[U-100% 13C; U-100% 15N]		1.5 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

LACS Plot; CA

Referencing offset: 0.14 ppm, Outliers: 1 Detail

LACS Plot; CB

Referencing offset: 0.14 ppm, Outliers: 1 Detail

LACS Plot; HA

Referencing offset: 0.09 ppm, Outliers: 1 Detail

Protein Blocks Logo

Calculated from 20 models in PDB: 2N66, Strand ID: A Detail

Release date

2016-08-14

Related entities 1. sweeter mutant (D40K) of sweet protein Brazzein, : 1 : 9 : 9 entities Detail

Experiments performed 10 experiments Detail

1 2D 1H-15N HSQC

Instrument

Varian INOVA - 600 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 310 K, pH 5.2

#	Name	Isotope labeling	Type	Concentration
1	D40K	[U-100% 13C; U-100% 15N]		1.5 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

2 3D CBCA(CO)NH

Instrument

Varian INOVA - 600 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 310 K, pH 5.2

#	Name	Isotope labeling	Type	Concentration
1	D40K	[U-100% 13C; U-100% 15N]		1.5 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

3 3D HNCO

Instrument

Varian INOVA - 600 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 310 K, pH 5.2

#	Name	Isotope labeling	Type	Concentration
1	D40K	[U-100% 13C; U-100% 15N]		1.5 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

4 3D HNCACB

Instrument

Varian INOVA - 600 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 310 K, pH 5.2

#	Name	Isotope labeling	Type	Concentration
1	D40K	[U-100% 13C; U-100% 15N]		1.5 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

5 3D HCCH-TOCSY

Instrument

Varian INOVA - 600 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 310 K, pH 5.2

#	Name	Isotope labeling	Type	Concentration
1	D40K	[U-100% 13C; U-100% 15N]		1.5 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

6 3D 1H-13C NOESY aliphatic

Instrument

Varian INOVA - 600 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 310 K, pH 5.2

#	Name	Isotope labeling	Type	Concentration
1	D40K	[U-100% 13C; U-100% 15N]		1.5 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

7 3D 1H-15N NOESY

Instrument

Varian INOVA - 600 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 310 K, pH 5.2

#	Name	Isotope labeling	Type	Concentration
1	D40K	[U-100% 13C; U-100% 15N]		1.5 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

8 2D 1H-13C HSQC

Instrument

Varian INOVA - 600 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 310 K, pH 5.2

#	Name	Isotope labeling	Type	Concentration
1	D40K	[U-100% 13C; U-100% 15N]		1.5 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

9 3D HCCH-COSY

Instrument

Varian INOVA - 600 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 310 K, pH 5.2

#	Name	Isotope labeling	Type	Concentration
1	D40K	[U-100% 13C; U-100% 15N]		1.5 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

10 3D 1H-13C NOESY aromatic

Instrument

Varian INOVA - 600 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 310 K, pH 5.2

#	Name	Isotope labeling	Type	Concentration
1	D40K	[U-100% 13C; U-100% 15N]		1.5 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

NMR combined restraints 4 contents Detail

Properties

Input source #1: NMR data (NEF) - Assigned chemical shifts, Distance restraints, Dihedral angle restraints	combined_25753_2n66.nef
Input source #2: Coordindates	2n66.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:52:CYS:SG	oxidized, CA 56.772, CB 41.6 ppm	oxidized, CA 54.8, CB 44.0 ppm	2.005
A:16:CYS:SG	A:37:CYS:SG	oxidized, CA 57.8, CB 40.254 ppm	oxidized, CA 56.2, CB 45.7 ppm	2.067
A:22:CYS:SG	A:47:CYS:SG	oxidized, CA 58.1, CB 38.5 ppm	oxidized, CA 54.7, CB 38.0 ppm	1.959
A:26:CYS:SG	A:49:CYS:SG	oxidized, CA 60.2, CB 36.6 ppm	oxidized, CA 51.387, CB 35.547 ppm	1.982

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--------50----
QDKCKKVYENYPVSKCQLANQCNYDCKLDKHARSGECFYKEKRNLQCICDYCEY
||||||||||||||||||||||||||||||||||||||||||||||||||||||
QDKCKKVYENYPVSKCQLANQCNYDCKLDKHARSGECFYKEKRNLQCICDYCEY

Chain assignments

Entity_assembly_ID (NMR)	Auth_asym_ID (model)	Length	Unmapped	Conflict	Sequence coverage (%)
A	A	54	0	0	100.0

Content subtype: combined_25753_2n66.nef

#	Content subtype	Saveframe	Status	# of rows (sets)	Experiment type	Sequence coverage (%)
1	Covalent bonds	assembly	OK	4		No information
1	Assigned chemical shifts	assigned_chem_shift_list_1	Warning	534		96.3 (chain: A, length: 54)
1	Distance restraints	DYANA/DIANA_distance_constraints_2	Warning	1341 (1)	noe	96.3 (chain: A, length: 54)
1	Dihedral angle restraints	DYANA/DIANA_dihedral_3	OK	64 (64)	.	87.0 (chain: A, length: 54)

Assigned chemical shifts

Saveframe: assigned_chem_shift_list_1
- Status: Warning
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 54, Sequence coverage: 96.3%
- Total number of assignments
  - ¹H chemical shifts: 321
  - ¹³C chemical shifts: 156
  - ¹⁵N chemical shifts: 57
- 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_2
- Status: Warning
- Experiment type: noe
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 54, Sequence coverage: 96.3%
- Total number of restraints: 1341
- Weight of restraints: 1.0 (1341)
- Potential type of restraints: upper-bound-parabolic (1341)
- Range of distance target values: 2.24, 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

Dihedral angle restraints

Saveframe: DYANA/DIANA_dihedral_3
- Status: OK
- Experiment type: .
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 54, Sequence coverage: 87.0%
- Total number of restraints: 64
- Total number of restraints per polymer type: 64
- Weight of restraints: 1.0 (64)
- Potential type of restraints: square-well-parabolic (64)
- Plot of Phi-Psi angle restraints
- Dihedral angle restraints per residue
  - Chain_ID: A

Keywords Brazzein, Sweet protein, Sweeter mutant D40K