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BMRB: 15995

2K9P

Structure of TM1_TM2 in LPPG micelles

Authors

Neumoin, A., Zerbe, O., Fred, N.

Assembly

TM1 TM2 in LPPG micelles

Entity

1. TM1 TM2 in LPPG micelles (polymer, Thiol state: not present), 80 monomers, 8751.091 Da Detail

GNGSTITFDE LQGLVNSTVT QAILFGVRSG AAALTLIVVW ITSRSRKTPI FIINQVSLFL IILHSALYFK YLLSNYSSVT

Formula weight

8751.091 Da

Source organism

Saccharomyces cerevisiae

Exptl. method

Refine. method

simulated annealing, TORSION ANGLE DYNAMICS

Data set

assigned_chemical_shifts

Chem. Shift Complete

Sequence coverage: 100.0 %, Completeness: 89.0 %, Completeness (bb): 96.7 % Detail

Polymer type: polypeptide(L)

	Total	¹H	¹³C	¹⁵N
All	89.0 % (837 of 940)	86.1 % (408 of 474)	92.6 % (351 of 379)	89.7 % (78 of 87)
Backbone	96.7 % (462 of 478)	97.6 % (160 of 164)	96.2 % (226 of 235)	96.2 % (76 of 79)
Sidechain	83.8 % (450 of 537)	80.0 % (248 of 310)	91.3 % (200 of 219)	25.0 % (2 of 8)
Aromatic	57.8 % (52 of 90)	57.8 % (26 of 45)	56.8 % (25 of 44)	100.0 % (1 of 1)
Methyl	98.5 % (132 of 134)	97.0 % (65 of 67)	100.0 % (67 of 67)

1. TM1 TM2 in LPPG micelles

GNGSTITFDE LQGLVNSTVT QAILFGVRSG AAALTLIVVW ITSRSRKTPI FIINQVSLFL IILHSALYFK YLLSNYSSVT

Show sample condition

Sample #1

Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for backbone assignment

#	Name	Isotope labeling	Type	Concentration
1	LPPG	natural abundance		200 mM
2	Na-phosphate buffer	natural abundance		20 mM
3	TM1-TM2 peptide	[U-100% 13C; U-100% 15N]		0.4 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

Sample #2

Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for 15N-NOESY measurement

#	Name	Isotope labeling	Type	Concentration
6	LPPG	natural abundance		200 mM
7	Na-phosphate buffer	natural abundance		20 mM
8	TM1-TM2 peptide	[U-100% 15N]		0.4 mM
9	H2O	natural abundance		90 %
10	D2O	natural abundance		10 %

Sample #3

Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for 15N-NOESY measurement on a 80%-deuterated background

#	Name	Isotope labeling	Type	Concentration
11	LPPG	natural abundance		200 mM
12	Na-phosphate buffer	natural abundance		20 mM
13	TM1-TM2 peptide	[U-100% 15N; 80% 2H]		0.4 mM
14	H2O	natural abundance		90 %
15	D2O	natural abundance		10 %

Sample #4

Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for 13C-NOESY measurement

#	Name	Isotope labeling	Type	Concentration
16	LPPG	[>85% 2H]		200 mM
17	Na-phosphate buffer	natural abundance		20 mM
18	TM1-TM2 peptide	[U-100% 13C; U-100% 15N]		0.4 mM
19	H2O	natural abundance		90 %
20	D2O	natural abundance		10 %

Sample #5

Solvent system 100% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details For 13C-NOESY measurement on selectively protonated Me-groups of Ile, Leu and Val residues on otherwise deuterated background.

#	Name	Isotope labeling	Type	Concentration
21	LPPG	[>85% 2H]		200 mM
22	Na-phosphate buffer	natural abundance		20 mM
23	TM1-TM2 peptide	[U-100% 13C; U-100% 15N; >98% 2H; Me(Ile(Hd1),Leu(Hd),Val(Hg)) 1H]		0.4 mM
24	D2O	natural abundance		100 %

Protein Blocks Logo

Calculated from 20 models in PDB: 2K9P, Strand ID: A Detail

Release date

2009-05-17

Citation

Structure of a double transmembrane fragment of a G-protein-coupled receptor in micelles
Neumoin, A., Cohen, L., Arshava, B., Tantry, S., Becker, J.M., Zerbe, O., Naider, F.
Biophys. J. (2009), 96, 3187-3196, PubMed 19383463 , DOI 10.1016/j.bpj.2009.01.012 , Abstract

Related entities 1. TM1 TM2 in LPPG micelles, : 1 : 4 entities Detail

Interaction partners 1. TM1 TM2 in LPPG micelles, : 8 interactors Detail

More details for 8 interactors identified by 6 citations

Experiments performed 13 experiments Detail

1 2D 1H-15N HSQC

Instrument

Bruker Avance - 700 MHz all tripleresonance experiments for assignment, 15N-NOESY, 15N-relaxation data

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for backbone assignment

#	Name	Isotope labeling	Type	Concentration
1	LPPG	natural abundance		200 mM
2	Na-phosphate buffer	natural abundance		20 mM
3	TM1-TM2 peptide	[U-100% 13C; U-100% 15N]		0.4 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

2 2D 1H-13C HSQC

Instrument

Bruker Avance - 700 MHz all tripleresonance experiments for assignment, 15N-NOESY, 15N-relaxation data

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for backbone assignment

#	Name	Isotope labeling	Type	Concentration
1	LPPG	natural abundance		200 mM
2	Na-phosphate buffer	natural abundance		20 mM
3	TM1-TM2 peptide	[U-100% 13C; U-100% 15N]		0.4 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

3 3D HNCO

Instrument

Bruker Avance - 700 MHz all tripleresonance experiments for assignment, 15N-NOESY, 15N-relaxation data

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for backbone assignment

#	Name	Isotope labeling	Type	Concentration
1	LPPG	natural abundance		200 mM
2	Na-phosphate buffer	natural abundance		20 mM
3	TM1-TM2 peptide	[U-100% 13C; U-100% 15N]		0.4 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

4 3D HNCA

Instrument

Bruker Avance - 700 MHz all tripleresonance experiments for assignment, 15N-NOESY, 15N-relaxation data

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for backbone assignment

#	Name	Isotope labeling	Type	Concentration
1	LPPG	natural abundance		200 mM
2	Na-phosphate buffer	natural abundance		20 mM
3	TM1-TM2 peptide	[U-100% 13C; U-100% 15N]		0.4 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

5 3D HNCACB

Instrument

Bruker Avance - 700 MHz all tripleresonance experiments for assignment, 15N-NOESY, 15N-relaxation data

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for backbone assignment

#	Name	Isotope labeling	Type	Concentration
1	LPPG	natural abundance		200 mM
2	Na-phosphate buffer	natural abundance		20 mM
3	TM1-TM2 peptide	[U-100% 13C; U-100% 15N]		0.4 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

6 3D HBHA(CO)NH

Instrument

Bruker Avance - 700 MHz all tripleresonance experiments for assignment, 15N-NOESY, 15N-relaxation data

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for backbone assignment

#	Name	Isotope labeling	Type	Concentration
1	LPPG	natural abundance		200 mM
2	Na-phosphate buffer	natural abundance		20 mM
3	TM1-TM2 peptide	[U-100% 13C; U-100% 15N]		0.4 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

7 3D CBCA(CO)NH

Instrument

Bruker Avance - 700 MHz all tripleresonance experiments for assignment, 15N-NOESY, 15N-relaxation data

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for backbone assignment

#	Name	Isotope labeling	Type	Concentration
1	LPPG	natural abundance		200 mM
2	Na-phosphate buffer	natural abundance		20 mM
3	TM1-TM2 peptide	[U-100% 13C; U-100% 15N]		0.4 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

8 3D HN(CO)CA

Instrument

Bruker Avance - 700 MHz all tripleresonance experiments for assignment, 15N-NOESY, 15N-relaxation data

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for backbone assignment

#	Name	Isotope labeling	Type	Concentration
1	LPPG	natural abundance		200 mM
2	Na-phosphate buffer	natural abundance		20 mM
3	TM1-TM2 peptide	[U-100% 13C; U-100% 15N]		0.4 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

9 3D HN(CA)CO

Instrument

Bruker Avance - 700 MHz all tripleresonance experiments for assignment, 15N-NOESY, 15N-relaxation data

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for backbone assignment

#	Name	Isotope labeling	Type	Concentration
1	LPPG	natural abundance		200 mM
2	Na-phosphate buffer	natural abundance		20 mM
3	TM1-TM2 peptide	[U-100% 13C; U-100% 15N]		0.4 mM
4	H2O	natural abundance		90 %
5	D2O	natural abundance		10 %

10 3D 1H-15N NOESY

Instrument

Bruker Avance - 700 MHz all tripleresonance experiments for assignment, 15N-NOESY, 15N-relaxation data

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for 15N-NOESY measurement

#	Name	Isotope labeling	Type	Concentration
6	LPPG	natural abundance		200 mM
7	Na-phosphate buffer	natural abundance		20 mM
8	TM1-TM2 peptide	[U-100% 15N]		0.4 mM
9	H2O	natural abundance		90 %
10	D2O	natural abundance		10 %

11 3D 1H-15N NOESY

Instrument

Bruker Avance - 700 MHz all tripleresonance experiments for assignment, 15N-NOESY, 15N-relaxation data

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for 15N-NOESY measurement on a 80%-deuterated background

#	Name	Isotope labeling	Type	Concentration
11	LPPG	natural abundance		200 mM
12	Na-phosphate buffer	natural abundance		20 mM
13	TM1-TM2 peptide	[U-100% 15N; 80% 2H]		0.4 mM
14	H2O	natural abundance		90 %
15	D2O	natural abundance		10 %

12 3D 1H-13C NOESY

Instrument

Bruker Avance - 900 MHz 13C-NOESY

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details for 13C-NOESY measurement

#	Name	Isotope labeling	Type	Concentration
16	LPPG	[>85% 2H]		200 mM
17	Na-phosphate buffer	natural abundance		20 mM
18	TM1-TM2 peptide	[U-100% 13C; U-100% 15N]		0.4 mM
19	H2O	natural abundance		90 %
20	D2O	natural abundance		10 %

13 3D 1H-13C NOESY

Instrument

Bruker Avance - 900 MHz 13C-NOESY

Sample

State isotropic, Solvent system 100% D2O, Pressure 1 atm, Temperature 320 K, pH 6.4, Details For 13C-NOESY measurement on selectively protonated Me-groups of Ile, Leu and Val residues on otherwise deuterated background.

#	Name	Isotope labeling	Type	Concentration
21	LPPG	[>85% 2H]		200 mM
22	Na-phosphate buffer	natural abundance		20 mM
23	TM1-TM2 peptide	[U-100% 13C; U-100% 15N; >98% 2H; Me(Ile(Hd1),Leu(Hd),Val(Hg)) 1H]		0.4 mM
24	D2O	natural abundance		100 %

NMR combined restraints 4 contents Detail

Properties

Input source #1: NMR data (NEF) - Assigned chemical shifts, Distance restraints, Dihedral angle restraints	combined_15995_2k9p.nef
Input source #2: Coordindates	2k9p.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

--------40--------50--------60--------70--------80--------90-------100-------110
GNGSTITFDELQGLVNSTVTQAILFGVRSGAAALTLIVVWITSRSRKTPIFIINQVSLFLIILHSALYFKYLLSNYSSVT
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
GNGSTITFDELQGLVNSTVTQAILFGVRSGAAALTLIVVWITSRSRKTPIFIINQVSLFLIILHSALYFKYLLSNYSSVT
--------10--------20--------30--------40--------50--------60--------70--------80

Chain assignments

Entity_assembly_ID (NMR)	Auth_asym_ID (model)	Length	Unmapped	Conflict	Sequence coverage (%)
A	A	80	0	0	100.0

Content subtype: combined_15995_2k9p.nef

#	Content subtype	Saveframe	Status	# of rows (sets)	Experiment type	Sequence coverage (%)
1	Assigned chemical shifts	assigned_chem_shift_list_1	Warning	829		100.0 (chain: A, length: 80)
1	Distance restraints	DYANA/DIANA_distance_constraints_2	Warning	1850 (1)	noe	96.3 (chain: A, length: 80)
1	Dihedral angle restraints	DYANA/DIANA_dihedral_4	OK	315 (315)	.	86.3 (chain: A, length: 80)

Assigned chemical shifts

Saveframe: assigned_chem_shift_list_1
- Status: Warning
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 80, Sequence coverage: 100.0%
- Total number of assignments
  - ¹H chemical shifts: 403
  - ¹³C chemical shifts: 350
  - ¹⁵N chemical shifts: 76
- 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: DYANA/DIANA_distance_constraints_2
- Status: Warning
- Experiment type: noe
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 80, Sequence coverage: 96.3%
- Total number of restraints: 1850
- Weight of restraints: 1.0 (1850)
- Potential type of restraints: upper-bound-parabolic (1850)
- Range of distance target values: 2.5, 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_4
- Status: OK
- Experiment type: .
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 80, Sequence coverage: 86.3%
- Total number of restraints: 315
- Total number of restraints per polymer type: 315
- Weight of restraints: 1.0 (315)
- Potential type of restraints: square-well-parabolic (315)
- Plot of Chi1-Chi2 angle restraints
- Dihedral angle restraints per residue
  - Chain_ID: A

Keywords fragment, GPCR, micelle, NMR, structurral biology