3O65

Crystal structure of a Josephin-ubiquitin complex: Evolutionary restraints on ataxin-3 deubiquitinating activity

X-RAY DIFFRACTION

Crystallization

Crystalization Experiments
ID	Method	pH	Temperature	Details
1		6.5	291	The protein complex was at 8 mg/ml in 10 mM Tris pH 7,5mm DTT. 2 microlitres drops were set up under paraffin oil. To 1.1 microlitres of precipitant (1.6 M sodium citrate pH 6.5, pH adjusted with HCl) 0.9 microlitres of protein was added, the final concentration of precipitant was therefore 0.88 M sodium citrate. For cryo crystals were passed through the 1.6 M sodium citrate pH 6.5 preciptant solution and then flash frozen in liquid nitrogen, microbatch under paraffin oil, temperature 291K

Crystal Properties
Matthews coefficient	Solvent content
4.33	71.59

Crystal Data

Unit Cell
Length ( Å )	Angle ( ˚ )
a = 159.15	α = 90
b = 159.15	β = 90
c = 146.29	γ = 120

Symmetry
Space Group	P 3 2 1

Diffraction

Diffraction Experiment
ID #	Crystal ID	Scattering Type	Data Collection Temperature	Detector	Detector Type	Details	Collection Date	Monochromator	Protocol
1	1	x-ray		CCD	ADSC QUANTUM 210	Toroidal focusing mirror	2008-10-10	M	SINGLE WAVELENGTH

Radiation Source
ID #	Source	Type	Wavelength List	Synchrotron Site	Beamline
1	SYNCHROTRON	NSLS BEAMLINE X6A	0.9780	NSLS	X6A

Data Collection

Overall
ID #	Resolution (High)	Resolution (Low)	Percent Possible (Observed)	R Merge 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	2.7		98.4	0.096	15.42			112055		-3	54.522

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)
	2.7	2.77	92.9		0.588	1.6

Refinement

Statistics
Diffraction ID	Structure Solution Method	Resolution (High)	Resolution (Low)	Cut-off Sigma (F)	Number Reflections (Observed)	Number Reflections (R-Free)	Percent Reflections (Observed)	R-Factor (Observed)	R-Work	R-Free	Mean Isotropic B
X-RAY DIFFRACTION	SAD	2.7	19.903	0.04	54826	2011	93.08	0.1781	0.1763	0.2242	78.9387

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]
1.9061			1.9061		-3.8123

RMS Deviations
Key	Refinement Restraint Deviation
f_dihedral_angle_d	15.888
f_angle_d	1.14
f_chiral_restr	0.074
f_bond_d	0.009
f_plane_restr	0.004

Non-Hydrogen Atoms Used in Refinement
Non-Hydrogen Atoms	Number
Protein Atoms	8343
Nucleic Acid Atoms
Solvent Atoms	152
Heterogen Atoms	1

Software

Software
Software Name	Purpose
XSCALE	data scaling
RESOLVE	phasing
PHENIX	refinement
PDB_EXTRACT	data extraction
HKL-2000	data collection
XDS	data reduction
PHENIX	phasing

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.