2KD1

Solution NMR structure of the integrase-like domain from Bacillus cereus ordered locus BC_1272. Northeast Structural Genomics Consortium Target BcR268F

SOLUTION NMR

NMR Experiment
Experiment	Type	Sample Contents	Solvent	Ionic Strength	pH	Pressure	Temperature (K)	Spectrometer
1	2D 1H-15N HSQC	0.986 mM [U-100% 13C; U-100% 15N] protein-1, 50 uM DSS-2, 200 mM sodium chloride-3, 20 mM MES-4, 10 mM DTT-5, 0.02 % NaN3-6, 5 mM Calcium Chloride-7	90% H2O/10% D2O	0.2	6.5	ambient	298
2	2D 1H-13C HSQC	0.986 mM [U-100% 13C; U-100% 15N] protein-1, 50 uM DSS-2, 200 mM sodium chloride-3, 20 mM MES-4, 10 mM DTT-5, 0.02 % NaN3-6, 5 mM Calcium Chloride-7	90% H2O/10% D2O	0.2	6.5	ambient	298
3	3D HNCO	0.986 mM [U-100% 13C; U-100% 15N] protein-1, 50 uM DSS-2, 200 mM sodium chloride-3, 20 mM MES-4, 10 mM DTT-5, 0.02 % NaN3-6, 5 mM Calcium Chloride-7	90% H2O/10% D2O	0.2	6.5	ambient	298
4	3D HNCA	0.986 mM [U-100% 13C; U-100% 15N] protein-1, 50 uM DSS-2, 200 mM sodium chloride-3, 20 mM MES-4, 10 mM DTT-5, 0.02 % NaN3-6, 5 mM Calcium Chloride-7	90% H2O/10% D2O	0.2	6.5	ambient	298
5	3D HNCACB	0.986 mM [U-100% 13C; U-100% 15N] protein-1, 50 uM DSS-2, 200 mM sodium chloride-3, 20 mM MES-4, 10 mM DTT-5, 0.02 % NaN3-6, 5 mM Calcium Chloride-7	90% H2O/10% D2O	0.2	6.5	ambient	298
6	3D CBCA(CO)NH	0.986 mM [U-100% 13C; U-100% 15N] protein-1, 50 uM DSS-2, 200 mM sodium chloride-3, 20 mM MES-4, 10 mM DTT-5, 0.02 % NaN3-6, 5 mM Calcium Chloride-7	90% H2O/10% D2O	0.2	6.5	ambient	298
7	3D HCCH-TOCSY	0.986 mM [U-100% 13C; U-100% 15N] protein-1, 50 uM DSS-2, 200 mM sodium chloride-3, 20 mM MES-4, 10 mM DTT-5, 0.02 % NaN3-6, 5 mM Calcium Chloride-7	90% H2O/10% D2O	0.2	6.5	ambient	298
8	3D 1H-13C NOESY	0.986 mM [U-100% 13C; U-100% 15N] protein-1, 50 uM DSS-2, 200 mM sodium chloride-3, 20 mM MES-4, 10 mM DTT-5, 0.02 % NaN3-6, 5 mM Calcium Chloride-7	90% H2O/10% D2O	0.2	6.5	ambient	298
9	3D 1H-15N NOESY	0.986 mM [U-100% 13C; U-100% 15N] protein-1, 50 uM DSS-2, 200 mM sodium chloride-3, 20 mM MES-4, 10 mM DTT-5, 0.02 % NaN3-6, 5 mM Calcium Chloride-7	90% H2O/10% D2O	0.2	6.5	ambient	298
10	3D HCCH-COSY	0.986 mM [U-100% 13C; U-100% 15N] protein-1, 50 uM DSS-2, 200 mM sodium chloride-3, 20 mM MES-4, 10 mM DTT-5, 0.02 % NaN3-6, 5 mM Calcium Chloride-7	90% H2O/10% D2O	0.2	6.5	ambient	298
11	3D CCH-TOCSY	0.986 mM [U-100% 13C; U-100% 15N] protein-1, 50 uM DSS-2, 200 mM sodium chloride-3, 20 mM MES-4, 10 mM DTT-5, 0.02 % NaN3-6, 5 mM Calcium Chloride-7	90% H2O/10% D2O	0.2	6.5	ambient	298
12	3D HBHA(CO)NH	0.986 mM [U-100% 13C; U-100% 15N] protein-1, 50 uM DSS-2, 200 mM sodium chloride-3, 20 mM MES-4, 10 mM DTT-5, 0.02 % NaN3-6, 5 mM Calcium Chloride-7	90% H2O/10% D2O	0.2	6.5	ambient	298
13	2D 1H-15N HSQC	0.953 mM [5% 13C; U-100% 15N] protein-8, 50 uM DSS-9, 200 mM sodium chloride-10, 20 mM MES-11, 10 mM DTT-12, 0.02 % NaN3-13, 5 mM Calcium Chloride-14	90% H2O/10% D2O	0.2	6.5	ambient	298
14	N15 T1	0.953 mM [5% 13C; U-100% 15N] protein-8, 50 uM DSS-9, 200 mM sodium chloride-10, 20 mM MES-11, 10 mM DTT-12, 0.02 % NaN3-13, 5 mM Calcium Chloride-14	90% H2O/10% D2O	0.2	6.5	ambient	298
15	N15 T2	0.953 mM [5% 13C; U-100% 15N] protein-8, 50 uM DSS-9, 200 mM sodium chloride-10, 20 mM MES-11, 10 mM DTT-12, 0.02 % NaN3-13, 5 mM Calcium Chloride-14	90% H2O/10% D2O	0.2	6.5	ambient	298
16	HET noe	0.953 mM [5% 13C; U-100% 15N] protein-8, 50 uM DSS-9, 200 mM sodium chloride-10, 20 mM MES-11, 10 mM DTT-12, 0.02 % NaN3-13, 5 mM Calcium Chloride-14	90% H2O/10% D2O	0.2	6.5	ambient	298
17	N15 trosy iso	0.953 mM [5% 13C; U-100% 15N] protein-8, 50 uM DSS-9, 200 mM sodium chloride-10, 20 mM MES-11, 10 mM DTT-12, 0.02 % NaN3-13, 5 mM Calcium Chloride-14	90% H2O/10% D2O	0.2	6.5	ambient	298
18	N15 trosy PolyAcrylamide Gel	0.953 mM [5% 13C; U-100% 15N] protein-8, 50 uM DSS-9, 200 mM sodium chloride-10, 20 mM MES-11, 10 mM DTT-12, 0.02 % NaN3-13, 5 mM Calcium Chloride-14	90% H2O/10% D2O	0.2	6.5	ambient	298
19	2D 1H-13C HSQC stereomethyls	0.953 mM [5% 13C; U-100% 15N] protein-8, 50 uM DSS-9, 200 mM sodium chloride-10, 20 mM MES-11, 10 mM DTT-12, 0.02 % NaN3-13, 5 mM Calcium Chloride-14	90% H2O/10% D2O	0.2	6.5	ambient	298

NMR Spectrometer Information
Spectrometer	Manufacturer	Model	Field Strength
1	Bruker	AVANCE	600
2	Varian	INOVA	600
3	Bruker	AVANCE	800

NMR Refinement
Method	Details	Software
torsion angle dynamics	MONOMER BY GEL FILTRATION CHROMATOGRAPHY/LIGHT SCATTERING AND BY NMR. T1/T2(CPMG) (MS) = 592.3/97.44, TAUC = 5.39 (NS). CONSISTENT WITH MOLECULAR WEIGHT. STRUCTURE DETERMINED BY TRIPLE RESONANCE NMR SPECTROSCOPY. NOESY ASSIGNMENTS BY CYANA-3.0 WITH RDC. 20 OF 100 STRUCTURES LOWEST TARGET FUNCTION SELECTED WITH CYANA-3.0. SELECTED MODELS ARE FURTHER REFINED USING CNS IN EXPLICIT WATER SHELL AND RDC SAMPLE ALIGNED IN POLYACRYLAMIDE GEL (NILGES PROTOCOL WITH PARAM19). ASSIGNMENT STATS (ALL RESIDUES INCLUDED): BACKBONE 93.88%, SIDECHAIN 85.19%, AROMATIC (SC) 92.00%, STEREOSPECIFIC VL METHYL ASSIGNMENT 100%, UNAMBIGUOUS SIDECHAIN NH2 100%. STRUCTURE BASED ON 2127 NOE, 246 DIHE, 105 RDC. MAX NOE VIOLATION: 0.41 A (1MODEL); MAX DIHE VIOLATION: 6.1 DEG. TWO TOTAL CLOSE CONTACTS PER 20 MODELS. STRUCTURE QUALITY FACTOR (PSVS 1.3): ORDERED RESIDUES RANGES: 7-22, 25-103 FOR [S(PHI)+S(PSI)] > 1.8. SECONDARY STRUCTURE - ALPHA HELICES: 8-23, 25-35, 39-42, 53-66, 70-89 RMSD (ANG): BACKBONE 0.4, ALL HEAVY ATOMS 0.7. RAMACHANDRAN DISTRIBUTION: 94.0/6.0/0.0/0.0. PROCHECK (PSI-PHI): 0.28/1.42 (RAW/Z), PROCHECK (ALL): 0.23/1.36 (RAW/Z), MOLPROBITY CLASH: 14.45/-0.95 (RAW/Z). RPF SCORES ALL ASSIGNED RESIDUES (FIT OF NOESY PEAKLISTS TO STRUCTURE): RECALL: 0.97, PRECISION: 0.85, F-MEASURE: 0.91, DP-SCORE: 0.82. RDC STATISTICS FROM CYANA-3.0. DA = -4.301 HZ, RHOM = 0.3925; CORR. COEFF: 0.947 +/- 0.002, Q-FACTOR: 32.55 +/- 0.405%. AFTER CNS WATER REFINEMENT WITH RDC PALES COMPUTED CORRELATION COEFF: 0.913 AND Q-FACTOR: 40.08% LOWEST ENERGY MODEL.	TopSpin

NMR Ensemble Information
Conformer Selection Criteria	target function
Conformers Calculated Total Number	100
Conformers Submitted Total Number	20
Representative Model	1 (lowest energy)

Computation: NMR Software
#	Classification	Version	Software Name	Author
1	collection	TopSpin	2.1	Bruker Biospin
2	processing	NMRPipe		Delaglio, Grzesiek, Vuister, Zhu, Pfeifer and Bax
3	data analysis	Sparky		Goddard
4	chemical shift assignment	PINE		Bahrami, Markley, Assadi, and Eghbalnia
5	structure solution	CYANA	3.0 beta	Guntert, Mumenthaler and Wuthrich
6	geometry optimization	TALOS		Cornilescu, Delaglio and Bax
7	refinement	CNS	1.2	Brunger, Adams, Clore, Gros, Nilges and Read
8	validation	PSVS		Bhattacharya and Montelione
9	visualization	MOLMOL		Koradi, Billeter and Wuthrich
10	validation	Procheck		Laskowski, MacArthur, Smith, Jones, Hutchinson, Morris, Moss and Tho
11	refinement	PdbStat	5.1	Tejero, Montelione
12	processing	MolProbity		Richardson

RCSB PDB Core Operations are funded by the U.S. National Science Foundation (DBI-2321666), the US Department of Energy (DE-SC0019749), and the National Cancer Institute, National Institute of Allergy and Infectious Diseases, and National Institute of General Medical Sciences of the National Institutes of Health under grant R01GM133198.

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2KD1

Solution NMR structure of the integrase-like domain from Bacillus cereus ordered locus BC_1272. Northeast Structural Genomics Consortium Target BcR268F