Evidence: isothermal titration calorimetry, The apo-form was obtained by complete refolding of protein-ligand complex. The addition of ligand (SAH or SAM) induced energy release observed in ITC ...Evidence: isothermal titration calorimetry, The apo-form was obtained by complete refolding of protein-ligand complex. The addition of ligand (SAH or SAM) induced energy release observed in ITC experiment. It indicate that we are dealing with apo-from of WBSCR27 protein., native gel electrophoresis, Native gel electrophoresis showed that WBSCR27 in our samples is represented by a monomer., gel filtration, The gel filtration experiment indicated that WBSCR27 in our samples is represented by a monomer., NMR relaxation study, NMR relaxation data showed that WBSCR27 in our samples is represented by a monomer.
Type
Name
Symmetry operation
Number
identity operation
1_555
1
Buried area
0 Å2
ΔGint
0 kcal/mol
Surface area
16480 Å2
Method
PISA
NMR ensembles
Data
Criteria
Number of conformers (submitted / calculated)
20 / 200
structures with the lowest energy
Representative
Model #1
closest to the average
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Components
#1: Protein
Methyltransferase-like27 / Williams-Beuren syndrome critical region protein 27 / Williams-Beuren syndrome critical region ...Williams-Beuren syndrome critical region protein 27 / Williams-Beuren syndrome critical region protein 27 isoform alpha / Williams-Beuren syndrome critical region protein 27 isoform beta
Mass: 25988.480 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Mus musculus (house mouse) / Gene: Mettl27, BC002286, Wbscr27 / Plasmid: pET30aTEV-WBSCR27 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q8BGM4
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Experimental details
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Experiment
Experiment
Method: SOLUTION NMR
NMR experiment
Conditions-ID
Experiment-ID
Solution-ID
Sample state
Spectrometer-ID
Type
1
1
1
isotropic
1
2D 1H-15N HSQC
1
2
4
isotropic
2
2D 1H-15N HSQC
1
3
3
isotropic
2
3D HNCO
1
4
3
isotropic
2
3D HN(CA)CO
1
5
3
isotropic
2
3DHN(CO)CA
1
8
3
isotropic
2
3D HNCA
1
7
3
isotropic
2
3DCBCA(CO)NH
1
6
3
isotropic
2
3D HN(CA)CB
1
9
3
isotropic
2
3DHBHA(CO)NH
1
10
3
isotropic
2
3DHNHAHB
1
11
3
isotropic
2
3D 1H-15N TOCSY
1
12
4
isotropic
2
3D (H)CCH-TOCSY
1
13
3
isotropic
2
3D 1H-13C NOESY
1
14
2
anisotropic
3
IPAP
1
17
5
isotropic
3
3D 15N-1H NOESY
1
16
1
isotropic
1
2D DQF-COSY
1
18
3
isotropic
2
3D 15N-1H NOESY
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Sample preparation
Details
Type
Solution-ID
Contents
Label
Solvent system
solution
1
0.8 mM [U-15N] WBSCR27, 10 mM DTT, 0.2 % w/v sodium azide, 50 mM sodium phosphate, 50 mM sodium chloride, 95% H2O/5% D2O
15N-apoWB27_1
95% H2O/5% D2O
solution
2
0.8 mM [U-15N] WBSCR27, 10 mM DTT, 0.2 % sodium azide, 50 mM sodium phosphate, 50 mM sodium chloride, 0.225 % v/v phage Pf1 stock solution, 95% H2O/5% D2O
15N-apoWB27_RDC
95% H2O/5% D2O
solution
3
0.8 mM [U-95% 13C; U-99% 15N] WBSCR27, 10 mM DTT, 0.2 % sodium azide, 50 mM sodium phosphate, 50 mM sodium chloride, 95% H2O/5% D2O
15N_13C-apoWB27_h2o
95% H2O/5% D2O
solution
4
0.3 mM [99% 13CG]-Thr [99% 13CE]-Met [100% 1HG]-Thr [100% 1HE]-Met [U-100% D] [U-98% 15N] WBSCR27, 5 mM DTT, 0.2 % sodium azide, 50 mM sodium phosphate, 50 mM sodium chloride, 100% D2O
15N_13C-apoWB27_d2o
100% D2O
solution
5
0.3 mM [99% 13CG]-Thr [99% 13CE]-Met [100% 1HG]-Thr [100% 1HE]-Met [U-100% D] [U-98% 15N] WBSCR27, 5 mM DTT, 0.2 % sodium azide, 50 mM sodium phosphate, 50 mM sodium chloride, 95% H2O/5% D2O