2HJ8

Solution NMR structure of the C-terminal domain of the interferon alpha-inducible ISG15 protein from Homo sapiens. Northeast Structural Genomics target HR2873B


SOLUTION NMR
NMR Experiment
ExperimentTypeSample ContentsSolventIonic StrengthpHPressureTemperature (K)Spectrometer
13D_15N-separated_NOESY0.81 mM U-13C,15N HR2873B, 50 mM ammonium citrate, 5 mM CaCl2, 1x protease inhibitor, 0.02% NaN3, pH 6.55% D2O / 95% H2O50 mM ammonium citrate, 5 mM CaCl26.5ambient293
23D_13C-separated_NOESY0.81 mM U-13C,15N HR2873B, 50 mM ammonium citrate, 5 mM CaCl2, 1x protease inhibitor, 0.02% NaN3, pH 6.55% D2O / 95% H2O50 mM ammonium citrate, 5 mM CaCl26.5ambient293
3HNHA0.81 mM U-13C,15N HR2873B, 50 mM ammonium citrate, 5 mM CaCl2, 1x protease inhibitor, 0.02% NaN3, pH 6.55% D2O / 95% H2O50 mM ammonium citrate, 5 mM CaCl26.5ambient293
4high resolution 2D CH-HSQC (for stereospecific assignment of Val/Leu methyls)0.9 mM 5%-13C,U-15N HR2873B, 50 mM ammonium citrate, 5 mM CaCl2, 1x protease inhibitor, 0.02% NaN3, pH 6.55% D2O / 95% H2O50 mM ammonium citrate, 5 mM CaCl26.5ambient293
52D 15N,1H heteronuclear NOE0.81 mM U-13C,15N HR2873B, 50 mM ammonium citrate, 5 mM CaCl2, 1x protease inhibitor, 0.02% NaN3, pH 6.55% D2O / 95% H2O50 mM ammonium citrate, 5 mM CaCl26.5ambient293
63D TR backbone expts0.81 mM U-13C,15N HR2873B, 50 mM ammonium citrate, 5 mM CaCl2, 1x protease inhibitor, 0.02% NaN3, pH 6.55% D2O / 95% H2O50 mM ammonium citrate, 5 mM CaCl26.5ambient293
73D HCCH-COSY, 3D HCCH-TOCSYs0.81 mM U-13C,15N HR2873B, 50 mM ammonium citrate, 5 mM CaCl2, 1x protease inhibitor, 0.02% NaN3, pH 6.55% D2O / 95% H2O50 mM ammonium citrate, 5 mM CaCl26.5ambient293
NMR Spectrometer Information
SpectrometerManufacturerModelField Strength
1BrukerAVANCE600
2VarianINOVA600
NMR Refinement
MethodDetailsSoftware
simulated annealingTHE STRUCTURES ARE BASED ON A TOTAL OF 976 CONFORMATIONALLY-RESTRICTING NOE-DERIVED DISTANCE CONSTRAINTS, 80 DIHEDRAL ANGLE CONSTRAINTS, AND 40 HYDROGEN BOND CONSTRAINTS (14.6 CONSTRAINTS PER RESIDUE, 4.8 LONG RANGE CONSTRAINTS PER RESIDUE, COMPUTED FOR RESIDUES 86 to 160 BY PSVS 1.3). STRUCTURE DETERMINATION WAS PERFORMED ITERATIVELY USING AUTOSTRUCTURE (XPLOR-NIH). AFTER A FINAL XPLOR CALCULATION USING THE CONSTRAINTS DERIVED FROM AUTOSTRUCTURE, THE 20 LOWEST ENERGY STRUCTURES OUT OF 100 WERE FURTHER REFINED BY RESTRAINED MOLECULAR DYANMICS/ENERGY MINIMIZATION IN EXPLICIT WATER (CNS). THE N-TERMINAL MET AND UNSTRUCTURED C-TERMINUS OF THE PROTEIN (LRGG + LEHHHHHH TAG) WERE INCLUDED IN ALL STRUCTURE CALCULATIONS BUT HAVE BEEN OMITTED FROM THIS DEPOSITION.XwinNMR
NMR Ensemble Information
Conformer Selection Criteriastructures with lowest conformational energy
Conformers Calculated Total Number100
Conformers Submitted Total Number20
Representative Model1 (lowest energy)
Additional NMR Experimental Information
DetailsTHE STRUCTURE WAS DETERMINED USING TRIPLE RESONANCE NMR SPECTROSCOPY. AUTOMATED BACKBONE ASSIGNMENTS WERE MADE USING AUTOASSIGN, AND THE SIDE CHAIN ASSIGNMENTS WERE COMPLETED MANUALLY. AUTOMATIC NOESY ASSIGNMENTS AS WELL AS DISTANCE AND HYDROGEN BOND CONSTRAINTS WERE DETERMINED USING AUTOSTRUCTURE. DIHEDRAL ANGLE CONSTRAINTS WERE DETERMINED USING HYPER. COMPLETENESS OF NMR ASSIGNMENTS (EXCLUDING N-TERMINAL MET AND C-TERMINAL HHHHHH): BACKBONE, 99.8%, SIDE CHAIN, 95.3%, AROMATICS, 100%, STEREOSPECIFIC METHYL, 95.2%. FINAL STRUCTURE QUALITY FACTORS (FOR RESIDUES 86 TO 160, PSVS 1.3), WHERE ORDERED RESIDUES [S(PHI) + S(PSI) > 1.8] COMPRISE 89-94,100-108,111-124,126-132,135-136,139-158: (A) RMSD (ORDERED RESIDUES): BB, 0.6, HEAVY ATOM, 1.1. (B) RAMACHANDRAN STATISTICS FOR ORDERED RESIDUES: MOST FAVORED, 86.7%, ADDITIONALLY ALLOWED, 12.7%, GENEROUSLY ALLOWED, 0.6%, DISALLOWED, 0.0%. (C) PROCHECK SCORES FOR ORDERED RESIDUES (RAW/Z-): PHI-PSI, -0.60/-2.05, ALL, -0.44/-2.60. (D) MOLPROBITY CLASH SCORE (RAW/Z-): 28.41/-3.35. (E) RPF SCORES FOR GOODNESS OF FIT TO NOESY DATA (ALL RESIDUES): RECALL, 0.978, PRECISION, 0.910, F-MEASURE, 0.943, DP-SCORE, 0.811.
Computation: NMR Software
#ClassificationVersionSoftware NameAuthor
1collectionXwinNMR3.5pl6Bruker
2collectionVNMR6.1CVarian
3data analysisAutoAssign2.2.1Zimmerman, Moseley, Montelione
4data analysisSparky3.110Goddard & Kneller
5refinementAutoStructure2.1.1Huang & Montelione
6refinementXPLOR-NIH2.11.2Clore et al
7processingNMRPipe2.3Delaglio et al
8data analysisPdbStat4.1Tejero & Montelione
9data analysisPSVS1.3Bhattacharya, Hang, Montelione
10refinementCNS1.1BRUNGER,ADAMS,CLORE,DELANO,GROS,GROSSE-KUNSTLEVE,JIANG,KUSZEWSKI,NILGES,PANNU,READ,RICE,SIMONSON,WARREN