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BMRB: 16913

2KXG

The solution structure of the squash aspartic acid proteinase inhibitor (SQAPI)

Authors

Headey, S.J., MacAskill, U.K., Wright, M., Claridge, J.K., Edwards, P.J.B., Farley, P.C., Christeller, J.T., Laing, W.A., Pascal, S.M.

Assembly

squash aspartic acid proteinase inhibitor

Entity

1. squash aspartic acid proteinase inhibitor (polymer, Thiol state: not present), 95 monomers, 10408.83 Da Detail

GPGPAIGEVI GISVNDPRVK EIAEFALKQH AEQNLILAGV DAGQIIKGIP HWDNYYNLIL SAKHSPHEFS KFYNVVVLEK ASDNSLKLVA FVPLF

Formula weight

10408.83 Da

Source organism

Cucurbita maxima

Exptl. method

Refine. method

torsion angle dynamics, simulated annealing, simulated annealing

Data set

assigned_chemical_shifts

Chem. Shift Complete

Sequence coverage: 100.0 %, Completeness: 94.6 %, Completeness (bb): 96.4 % Detail

Polymer type: polypeptide(L)

	Total	¹H	¹³C
All	94.6 % (968 of 1023)	94.0 % (544 of 579)	95.5 % (424 of 444)
Backbone	96.4 % (452 of 469)	99.0 % (189 of 191)	94.6 % (263 of 278)
Sidechain	94.1 % (604 of 642)	91.5 % (355 of 388)	98.0 % (249 of 254)
Aromatic	89.1 % (90 of 101)	88.2 % (45 of 51)	90.0 % (45 of 50)
Methyl	97.6 % (123 of 126)	95.2 % (60 of 63)	100.0 % (63 of 63)

1. SQAPI

GPGPAIGEVI GISVNDPRVK EIAEFALKQH AEQNLILAGV DAGQIIKGIP HWDNYYNLIL SAKHSPHEFS KFYNVVVLEK ASDNSLKLVA FVPLF

Show sample condition

Sample

Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

LACS Plot; CA

Referencing offset: -0.28 ppm, Outliers: 1 Detail

LACS Plot; CB

Referencing offset: -0.28 ppm, Outliers: 1 Detail

LACS Plot; HA

Referencing offset: -0.04 ppm, Outliers: 1 Detail

LACS Plot; CO

Referencing offset: 0.71 ppm, Outliers: 3 Detail

Protein Blocks Logo

Calculated from 10 models in PDB: 2KXG, Strand ID: A Detail

Release date

2010-07-25

Citation

Solution structure of the squash aspartic acid proteinase inhibitor (SQAPI) and mutational analysis of pepsin inhibition
Headey, S.J., Macaskill, U.K., Wright, M.A., Claridge, J.K., Edwards, P.J.B., Farley, P.C., Christeller, J.T., Laing, W.A., Pascal, S.M.
J. Biol. Chem. (2010), 285, 27019-27025, PubMed 20538608 , DOI 10.1074/jbc.M110.137018 , Abstract

Related entities 1. squash aspartic acid proteinase inhibitor, : 1 : 25 entities Detail

Experiments performed 15 experiments Detail

1 2D 1H-15N HSQC

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

2 2D 1H-13C HSQC

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

3 2D 1H-1H NOESY

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

4 3D HNCACB

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

5 3D HNCA

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

6 3D HNCO

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

7 3D HN(CA)CO

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

8 3D HN(CO)CACB

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

9 3D HCCCONH TOCSY

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

10 3D HBHA(CO)NH

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

11 3D HCCH-TOCSY

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

12 2D (HB)CB(CGCD)HD

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

13 2D (HB)CB(CGCDCE)HE

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

14 3D 1H-13C NOESYHSQC

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

15 3D 1H-15N NOESYHSQC

Instrument

Bruker Avance - 700 MHz cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 323 K, pH 3

#	Name	Isotope labeling	Type	Concentration
1	SQAPI	[U-98% 13C; U-98% 15N]		0.6 mM
2	H2O	natural abundance		90 %
3	D2O	natural abundance		10 %

NMR combined restraints 5 contents Detail

Properties

Input source #1: NMR data (NEF) - Assigned chemical shifts, Distance restraints, Dihedral angle restraints	combined_16913_2kxg.nef
Input source #2: Coordindates	2kxg.cif
Diamagnetism of the molecular assembly	True (excluding Oxygen atoms)
Whether the assembly has a disulfide bond	None
Whether the assembly has a other bond	None
Whether the assembly contains a cyclic polymer	None
Overall data processing status	Warning

Disulfide bonds

None

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--------60--------70--------80--------90-----
GPGPAIGEVIGISVNDPRVKEIAEFALKQHAEQNLILAGVDAGQIIKGIPHWDNYYNLILSAKHSPHEFSKFYNVVVLEKASDNSLKLVAFVPLF
|||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
GPGPAIGEVIGISVNDPRVKEIAEFALKQHAEQNLILAGVDAGQIIKGIPHWDNYYNLILSAKHSPHEFSKFYNVVVLEKASDNSLKLVAFVPLF

Chain assignments

Entity_assembly_ID (NMR)	Auth_asym_ID (model)	Length	Unmapped	Conflict	Sequence coverage (%)
A	A	95	0	0	100.0

Content subtype: combined_16913_2kxg.nef

#	Content subtype	Saveframe	Status	# of rows (sets)	Experiment type	Sequence coverage (%)
1	Assigned chemical shifts	assigned_chem_shift_list_1	Warning	972		100.0 (chain: A, length: 95)
1	Distance restraints	CNS/XPLOR_distance_constraints_2	OK	1669 (1)	noe	98.9 (chain: A, length: 95)
2	Distance restraints	CNS/XPLOR_distance_constraints_4	OK	78 (1)	hbond	51.6 (chain: A, length: 95)
1	Dihedral angle restraints	CNS/XPLOR_dihedral_3	OK	137 (137)	.	93.7 (chain: A, length: 95)

Assigned chemical shifts

Saveframe: assigned_chem_shift_list_1
- Status: Warning
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 95, Sequence coverage: 100.0%
- Total number of assignments
  - ¹H chemical shifts: 551
  - ¹³C chemical shifts: 421
- Completeness of assignments
  - Entity_assemble_ID: A
- Peptide bond of PRO
- Tautomeric state of HIS
- 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

Distance restraints

Saveframe: CNS/XPLOR_distance_constraints_2
- Status: OK
- Experiment type: noe
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 95, Sequence coverage: 98.9%
- Total number of restraints: 1668
- Weight of restraints: 1.0 (1668)
- Potential type of restraints: square-well-parabolic (1668)
- Range of distance target values: 2.1, 3.65 (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: CNS/XPLOR_distance_constraints_4
  - Status: OK
  - Experiment type: hbond
  - Sequence coverage of experimental data
    - Entity_assembly_ID: A, Chain length: 95, Sequence coverage: 51.6%
  - Total number of restraints: 78
  - Weight of restraints: 1.0 (78)
  - Potential type of restraints: square-well-parabolic (78)
  - Range of distance target values: 1.85, 2.75 (Å)
  - Histogram of distance target values
  - Distance restraints per residue
    - Chain_ID: A
  - Distance restraints on contact map

Dihedral angle restraints

Saveframe: CNS/XPLOR_dihedral_3
- Status: OK
- Experiment type: .
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 95, Sequence coverage: 93.7%
- Total number of restraints: 137
- Total number of restraints per polymer type: 137
- Weight of restraints: 1.0 (137)
- Potential type of restraints: square-well-parabolic (137)
- Plot of Phi-Psi angle restraints
- Dihedral angle restraints per residue
  - Chain_ID: A

Keywords aspartic protease inhibitor, protein, SQAPI