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BMRB: 30324

5WOW

Solution NMR structure of cyclotide MCoTI-I

Authors

Schroeder, C.I., Kwon, S.

Assembly

Two inhibitor peptide topologies 2

Entity

1. Two inhibitor peptide topologies 2 (polymer, Thiol state: not present), 39 monomers, 4242.801 Da Detail

GGVCPKILQR CRRDSDCPGA CICRGNGYCG YPYDVPDYA

Formula weight

4242.801 Da

Source organism

Momordica cochinchinensis

Exptl. method

Data set

assigned_chemical_shifts

Chem. Shift Complete

Sequence coverage: 100.0 %, Completeness: 85.6 %, Completeness (bb): 83.2 % Detail

Polymer type: polypeptide(L)

	Total	¹H	¹³C	¹⁵N
All	85.6 % (356 of 416)	97.3 % (213 of 219)	66.9 % (107 of 160)	97.3 % (36 of 37)
Backbone	83.2 % (188 of 226)	100.0 % (80 of 80)	66.7 % (74 of 111)	97.1 % (34 of 35)
Sidechain	90.1 % (201 of 223)	95.7 % (133 of 139)	80.5 % (66 of 82)	100.0 % (2 of 2)
Aromatic	50.0 % (16 of 32)	100.0 % (16 of 16)	0.0 % (0 of 16)
Methyl	100.0 % (24 of 24)	100.0 % (12 of 12)	100.0 % (12 of 12)

1. Two inhibitor peptide topologies 2

GGVCPKILQR CRRDSDCPGA CICRGNGYCG YPYDVPDYA

Show sample condition

Sample #1

Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 298 K, pH 3.0, Details 0.5 mM MCoTI-HA, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	MCoTI-HA	natural abundance		0.5 mM

Sample #2

Solvent system 100% D2O, Pressure 1 atm, Temperature 298 K, pH 3.0, Details 0.5 mM MCoTI-HA, 100% D2O

#	Name	Isotope labeling	Type	Concentration
2	MCoTI-HA	natural abundance		0.5 mM

Release date

2017-08-17

Citation

Targeted Delivery of Cyclotides via Conjugation to a Nanobody
Kwon, S., Duarte, J.N., Li, Z., Ling, J.J., Cheneval, O., Durek, T., Schroeder, C.I., Craik, D.J., Ploegh, H.L.
ACS Chem. Biol. (2018), 13, 2973-2980, PubMed 30248263 , DOI 10.1021/acschembio.8b00653 , Abstract

Entries sharing articles BMRB: 1 entries Detail

BMRB: 30323 released on 2017-08-17
Title Solution NMR structure of cyclotide MCoTI-I

Related entities 1. Two inhibitor peptide topologies 2, : 1 : 11 entities Detail

Experiments performed 8 experiments Detail

1 2D 1H-1H TOCSY

Instrument

Bruker Avance - 600 MHz equipped with a cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 298 K, pH 3.0, Details 0.5 mM MCoTI-HA, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	MCoTI-HA	natural abundance		0.5 mM

2 2D 1H-1H NOESY

Instrument

Bruker Avance - 600 MHz equipped with a cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 298 K, pH 3.0, Details 0.5 mM MCoTI-HA, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	MCoTI-HA	natural abundance		0.5 mM

3 2D 1H-15N HSQC

Instrument

Bruker Avance - 600 MHz equipped with a cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 298 K, pH 3.0, Details 0.5 mM MCoTI-HA, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	MCoTI-HA	natural abundance		0.5 mM

4 2D DQF-COSY

Instrument

Bruker Avance - 600 MHz equipped with a cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 298 K, pH 3.0, Details 0.5 mM MCoTI-HA, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	MCoTI-HA	natural abundance		0.5 mM

5 2D 1H-13C HSQC

Instrument

Bruker Avance - 600 MHz equipped with a cryoprobe

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 298 K, pH 3.0, Details 0.5 mM MCoTI-HA, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	MCoTI-HA	natural abundance		0.5 mM

6 2D 1H-1H E COSY

Instrument

Bruker Avance - 600 MHz equipped with a cryoprobe

Sample

State isotropic, Solvent system 100% D2O, Pressure 1 atm, Temperature 298 K, pH 3.0, Details 0.5 mM MCoTI-HA, 100% D2O

#	Name	Isotope labeling	Type	Concentration
2	MCoTI-HA	natural abundance		0.5 mM

7 2D 1H-13C HSQC

Instrument

Bruker Avance - 600 MHz equipped with a cryoprobe

Sample

State isotropic, Solvent system 100% D2O, Pressure 1 atm, Temperature 298 K, pH 3.0, Details 0.5 mM MCoTI-HA, 100% D2O

#	Name	Isotope labeling	Type	Concentration
2	MCoTI-HA	natural abundance		0.5 mM

8 2D 1H-1H TOCSY

Instrument

Bruker Avance - 600 MHz equipped with a cryoprobe

Sample

State isotropic, Solvent system 100% D2O, Pressure 1 atm, Temperature 298 K, pH 3.0, Details 0.5 mM MCoTI-HA, 100% D2O

#	Name	Isotope labeling	Type	Concentration
2	MCoTI-HA	natural abundance		0.5 mM

NMR combined restraints 5 contents Detail

Properties

Input source #1: NMR data (NEF) - Assigned chemical shifts, Distance restraints, Dihedral angle restraints	combined_30324_5wow.nef
Input source #2: Coordindates	5wow.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	True (see coodinates for details)
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:21:CYS:SG	oxidized, CA 55.288, CB 42.173 ppm	oxidized, CA 57.511, CB 45.153 ppm	2.015
A:11:CYS:SG	A:23:CYS:SG	oxidized, CA 56.552, CB 50.486 ppm	oxidized, CA 57.592, CB 40.663 ppm	2.034
A:17:CYS:SG	A:29:CYS:SG	oxidized, CA 54.572, CB 42.797 ppm	oxidized, CA 58.039, CB 43.201 ppm	2.035

Other bonds (neither disulfide, covalent nor hydrogen bonds, e.g. Zinc–sulphur bond)

Ptnr_site_1	Ptnr_site_2	Redox_state_prediction_1	Redox_state_prediction_2	Distance (Å)
1:39:ALA:C	1:1:GLY:N	unknown	unknown	n/a

Non-standard residues

None

Sequence alignments

Entity_assembly_ID (NMR): A vs Auth_asym_ID (model): A

--------10--------20--------30---------
GGVCPKILQRCRRDSDCPGACICRGNGYCGYPYDVPDYA
|||||||||||||||||||||||||||||||||||||||
GGVCPKILQRCRRDSDCPGACICRGNGYCGYPYDVPDYA

Chain assignments

Entity_assembly_ID (NMR)	Auth_asym_ID (model)	Length	Unmapped	Conflict	Sequence coverage (%)
A	A	39	0	0	100.0

Content subtype: combined_30324_5wow.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_chemical_shifts_1	Warning	357		100.0 (chain: A, length: 39)
1	Distance restraints	CNS/XPLOR_distance_constraints_2	Warning	406 (1)	noe	97.4 (chain: A, length: 39)
2	Distance restraints	CNS/XPLOR_distance_constraints_4	Warning	12 (1)	hbond	28.2 (chain: A, length: 39)
1	Dihedral angle restraints	CNS/XPLOR_dihedral_3	OK	53 (53)	.	97.4 (chain: A, length: 39)

Assigned chemical shifts

Saveframe: assigned_chemical_shifts_1
- Status: Warning
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 39, Sequence coverage: 100.0%
- Total number of assignments
  - ¹H chemical shifts: 212
  - ¹³C chemical shifts: 105
  - ¹⁵N chemical shifts: 40
- 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: CNS/XPLOR_distance_constraints_2
- Status: Warning
- Experiment type: noe
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 39, Sequence coverage: 97.4%
- Total number of restraints: 406
- Weight of restraints: 1.0 (406)
- Potential type of restraints: square-well-parabolic (406)
- Range of distance target values: 1.4, 4.33 (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: Warning
  - Experiment type: hbond
  - Sequence coverage of experimental data
    - Entity_assembly_ID: A, Chain length: 39, Sequence coverage: 28.2%
  - Total number of restraints: 12
  - Weight of restraints: 1.0 (12)
  - Potential type of restraints: square-well-parabolic (12)
  - Range of distance target values: 1.0, 1.5 (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: CNS/XPLOR_dihedral_3
- Status: OK
- Experiment type: .
- Sequence coverage of experimental data
  - Entity_assembly_ID: A, Chain length: 39, Sequence coverage: 97.4%
- Total number of restraints: 53
- Total number of restraints per polymer type: 53
- Weight of restraints: 1.0 (53)
- Potential type of restraints: square-well-parabolic (53)
- Plot of Phi-Psi angle restraints
- Dihedral angle restraints per residue
  - Chain_ID: A

Keywords DE NOVO PROTEIN, bioactive epitope, cyclic cystine knot, cyclotide, grafted peptide, plant peptide