4UN4

Crystal structure of Cas9 bound to PAM-containing DNA target with mismatches at positions 1-2

X-RAY DIFFRACTION

Crystallization

Crystalization Experiments
ID	Method	pH	Temperature	Details
1		8.5		0.1 M TRIS-ACETATE, PH 8.5, 0.3 M KSCN, 15% (W/V) PEG 3350

Crystal Properties
Matthews coefficient	Solvent content
2.68	54.1

Crystal Data

Unit Cell
Length ( Å )	Angle ( ˚ )
a = 177.78	α = 90
b = 66.816	β = 111.39
c = 186.834	γ = 90

Symmetry
Space Group	C 1 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	100	PIXEL	DECTRIS PILATUS		2014-05-10	M	SINGLE WAVELENGTH

Radiation Source
ID #	Source	Type	Wavelength List	Synchrotron Site	Beamline
1	SYNCHROTRON	SLS BEAMLINE X06DA		SLS	X06DA

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	2.37	47.49	99.1	0.1	16.5	6.8		160249		2

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	2.37	2.46	91.5		0.81	2.2	6.3

Refinement

Statistics
Diffraction ID	Structure Solution Method	Starting model	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	MOLECULAR REPLACEMENT	PDB ENTRY 4UN3	2.371	47.493	1.3	160249	8029	98.72	0.2185	0.2169	0.2481

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]

RMS Deviations
Key	Refinement Restraint Deviation
f_dihedral_angle_d	15.884
f_angle_d	0.694
f_chiral_restr	0.045
f_bond_d	0.013
f_plane_restr	0.002

Non-Hydrogen Atoms Used in Refinement
Non-Hydrogen Atoms	Number
Protein Atoms	10716
Nucleic Acid Atoms	2503
Solvent Atoms	687
Heterogen Atoms	8

Software

Software
Software Name	Purpose
PHENIX	refinement
XDS	data reduction
SCALA	data scaling
PHASER	phasing

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.