5BXY

Crystal structure of RNA methyltransferase from Salinibacter ruber in complex with S-Adenosyl-L-homocysteine

X-RAY DIFFRACTION

Crystallization

Crystalization Experiments
ID	Method	pH	Temperature	Details
1	VAPOR DIFFUSION, SITTING DROP	8.5	289	0.2 ul of 15 mg/ml protein in 20 mM HEPES pH 7.5, 150 mM NaCl, 10% Glycerol, 0.1% Sodium Azide and 0.5 mM TCEP were mixed with 0.2 ul of the MCSG Suite 1 #23 (0.1M Tris, 20%w/v PEG 8K, 0.2M Magnesium Chloride, 6-Hydrate pH=8.5) and equilibrated against 1.5 M NaCl solution in 96 Well 3 drop Crystallization Plate (Swissci). Before crystallization protein was incubated with 1/50 v/v of 2 mg/ml chymotrypsin solution at 289 K for 3 hours

Crystal Properties
Matthews coefficient	Solvent content
2.27	45.81

Crystal Data

Unit Cell
Length ( Å )	Angle ( ˚ )
a = 43.574	α = 90
b = 86.04	β = 90
c = 85.057	γ = 90

Symmetry
Space Group	P 21 21 21

Diffraction

Diffraction Experiment
ID #	Crystal ID	Scattering Type	Data Collection Temperature	Detector	Detector Type	Details	Collection Date	Monochromator	Protocol
1	1	x-ray	100	CCD	MARMOSAIC 300 mm CCD		2015-04-16	M	SINGLE WAVELENGTH

Radiation Source
ID #	Source	Type	Wavelength List	Synchrotron Site	Beamline
1	SYNCHROTRON	APS BEAMLINE 21-ID-G	0.979	APS	21-ID-G

Data Collection

Overall
ID #	Resolution (High)	Resolution (Low)	Percent Possible (Observed)	R Merge I (Observed)	R Sym I (Observed)	Net I Over Average Sigma (I)	Redundancy	Number Reflections (All)	Number Reflections (Observed)	Observed Criterion Sigma (F)	Observed Criterion Sigma (I)	B (Isotropic) From Wilson Plot
1	1.75	50	94.8	0.099	0.099	17.8	5.1		31176		-3

Highest Resolution Shell
ID #	Resolution (High)	Resolution (Low)	Percent Possible (All)	Percent Possible (Observed)	R Merge I (Observed)	Mean I Over Sigma (Observed)	Redundancy	Number Unique Reflections (All)
1	1.75	1.78	98.4		0.497	2.2	4.7

Refinement

Statistics
Diffraction ID	Structure Solution Method	Cross Validation method	Resolution (High)	Resolution (Low)	Number Reflections (Observed)	Number Reflections (R-Free)	Percent Reflections (Observed)	R-Factor (Observed)	R-Work	R-Free	R-Free Selection Details	Mean Isotropic B
X-RAY DIFFRACTION	SAD	THROUGHOUT	1.75	50	29629	1508	94.61	0.1503	0.1484	0.1882	RANDOM	20.852

Temperature Factor Modeling
Anisotropic B[1][1]	Anisotropic B[1][2]	Anisotropic B[1][3]	Anisotropic B[2][2]	Anisotropic B[2][3]	Anisotropic B[3][3]
			0.13		-0.13

RMS Deviations
Key	Refinement Restraint Deviation
r_dihedral_angle_2_deg	29.87
r_dihedral_angle_4_deg	18.814
r_dihedral_angle_3_deg	11.765
r_dihedral_angle_1_deg	6.292
r_mcangle_it	4.481
r_mcbond_it	4.007
r_mcbond_other	3.98
r_angle_refined_deg	1.598
r_angle_other_deg	1.206
r_chiral_restr	0.089

RMS Deviations
Key	Refinement Restraint Deviation
r_dihedral_angle_2_deg	29.87
r_dihedral_angle_4_deg	18.814
r_dihedral_angle_3_deg	11.765
r_dihedral_angle_1_deg	6.292
r_mcangle_it	4.481
r_mcbond_it	4.007
r_mcbond_other	3.98
r_angle_refined_deg	1.598
r_angle_other_deg	1.206
r_chiral_restr	0.089
r_bond_refined_d	0.01
r_gen_planes_refined	0.007
r_bond_other_d	0.005
r_gen_planes_other	0.004

Non-Hydrogen Atoms Used in Refinement
Non-Hydrogen Atoms	Number
Protein Atoms	2348
Nucleic Acid Atoms
Solvent Atoms	341
Heterogen Atoms	56

Software

Software
Software Name	Purpose
SBC-Collect	data collection
HKL-2000	data scaling
SCALEPACK	data scaling
HKL-3000	data reduction
MLPHARE	phasing
HKL-3000	phasing
DM	phasing
SHELX	phasing
PDB_EXTRACT	data extraction
REFMAC	refinement
Coot	model building

RCSB PDB Core Operations are funded by the National Science Foundation (DBI-1832184), 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.