3THP

Crystal structure and RNA binding properties of the RRM/AlkB domains in human ABH8, an enzyme catalyzing tRNA hypermodification, Northeast Structural Genomics Consortium Target HR5601B

X-RAY DIFFRACTION

Starting Model(s)

Initial Refinement Model(s)
Type	Source	Accession Code	Details
experimental model	PDB	2CQ2	PDB entry 2CQ2 for RRM domain, PDB entry 2FDI for AlkB domain.
experimental model	PDB	2FDI	PDB entry 2CQ2 for RRM domain, PDB entry 2FDI for AlkB domain.

Crystallization

Crystalization Experiments
ID	Method	pH	Temperature	Details
1	microbatch under oil	7	293	30% PEG 3350 and 0.15 M D/L malic acid, 2.8 mM MnCl2, and 8.6 mM 2-oxyglutarate., pH 7, microbatch under oil, temperature 293K

Crystal Properties
Matthews coefficient	Solvent content
2.54	51.61

Crystal Data

Unit Cell
Length ( Å )	Angle ( ˚ )
a = 68.276	α = 90
b = 81.675	β = 90
c = 144.72	γ = 90

Symmetry
Space Group	I 2 2 2

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	MAR CCD 165 mm	mirrors	2010-11-10	M	SINGLE WAVELENGTH

Radiation Source
ID #	Source	Type	Wavelength List	Synchrotron Site	Beamline
1	SYNCHROTRON	NSLS BEAMLINE X4C	0.97885	NSLS	X4C

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	3.2	40	99.9	0.154	0.124	14.3	7.4	12782	12770			42.5

Highest Resolution Shell
ID #	Resolution (High)	Resolution (Low)	Percent Possible (All)	Percent Possible (Observed)	R Merge I (Observed)	R-Sym I (Observed)	Mean I Over Sigma (Observed)	Redundancy	Number Unique Reflections (All)
1	3.2	3.31	99.6		0.437	0.393	3.7	6.4	1293

Refinement

Statistics
Diffraction ID	Structure Solution Method	Cross Validation method	Starting model	Resolution (High)	Resolution (Low)	Number Reflections (All)	Number Reflections (Observed)	Number Reflections (R-Free)	Percent Reflections (Observed)	R-Factor (All)	R-Factor (Observed)	R-Work	R-Free	R-Free Selection Details	Mean Isotropic B
X-RAY DIFFRACTION	MOLECULAR REPLACEMENT	THROUGHOUT	PDB entry 2CQ2 for RRM domain, PDB entry 2FDI for AlkB domain.	3.2	35.57	12782	12202	1210	94.9	0.22	0.217	0.217	0.283	RANDOM	44.2

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]
-12.99			24.43		-11.44

RMS Deviations
Key	Refinement Restraint Deviation
c_dihedral_angle_d	25
c_angle_deg	1.1
c_improper_angle_d	0.76
c_bond_d	0.009

Non-Hydrogen Atoms Used in Refinement
Non-Hydrogen Atoms	Number
Protein Atoms	2413
Nucleic Acid Atoms
Solvent Atoms
Heterogen Atoms	12

Software

Software
Software Name	Purpose
ADSC	data collection
PHASER	phasing
CNS	refinement
DENZO	data reduction
SCALEPACK	data scaling

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 R01GM157729.