4X9W

PLK-1 polo-box domain in complex with Bioactive Imidazolium-containing phosphopeptide macrocycle 4C

X-RAY DIFFRACTION

Crystallization

Crystalization Experiments
ID	Method	pH	Temperature	Details
1	VAPOR DIFFUSION	8	291	Frozen stocks of protein at 37 mg/mL in 10 mM TRIS pH8, 0.5 M NaCl, 10 mM DTT were thawed and diluted to 10 mg/ml with the same buffer. Complexes with each of three macrocycle compounds (3b, 3c and 4b) were prepared by adding 100 mM stocks of the macrocycle in DMSO directly to the diluted protein to achieve a final concentration of 1 mM. Crystals were grown by hanging drop vapor diffusion, with drops made by mixing equal volumes of protein-macrocycle complex and well solution containing 2-6% PEG-3350. Crystals were cryo-protected by quickly dipping in a solution of 37.5% ethylene glycol in well solution and frozen in liquid nitrogen.

Crystal Properties
Matthews coefficient	Solvent content
1.92	36.02

Crystal Data

Unit Cell
Length ( Å )	Angle ( ˚ )
a = 36.052	α = 90
b = 51.525	β = 100.55
c = 57.449	γ = 90

Symmetry
Space Group	P 1 21 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	PSI PILATUS 6M		2013-11-30	M	SINGLE WAVELENGTH

Radiation Source
ID #	Source	Type	Wavelength List	Synchrotron Site	Beamline
1	SYNCHROTRON	APS BEAMLINE 24-ID-C	0.97950	APS	24-ID-C

Data Collection

Overall
ID #	Resolution (High)	Resolution (Low)	Percent Possible (Observed)	R Merge I (Observed)	Rrim I (All)	Rpim I (All)	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	1.798	100	95.7	0.067	0.073	0.028	7	6.3		18528			27.05

Highest Resolution Shell
ID #	Resolution (High)	Resolution (Low)	Percent Possible (All)	Percent Possible (Observed)	R Merge I (Observed)	Rrim I (All)	Rpim I (All)	CC (Half)	Mean I Over Sigma (Observed)	Redundancy	Number Unique Reflections (All)
1	1.798	1.83	80.2		0.301	0.329	0.132	0.965		5.4	753

Refinement

Statistics
Diffraction ID	Structure Solution Method	Cross Validation 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	R-Free Selection Details	Mean Isotropic B
X-RAY DIFFRACTION	MOLECULAR REPLACEMENT	FREE R-VALUE	3RQ7	1.798	38.065	1.36	18493	1850	95.42	0.1987	0.1944	0.2388	random

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	12.639
f_angle_d	0.846
f_chiral_restr	0.041
f_bond_d	0.004
f_plane_restr	0.003

Non-Hydrogen Atoms Used in Refinement
Non-Hydrogen Atoms	Number
Protein Atoms	1831
Nucleic Acid Atoms
Solvent Atoms	73
Heterogen Atoms	62

Software

Software
Software Name	Purpose
PHENIX	refinement
HKL-2000	data reduction
PDB_EXTRACT	data extraction
HKL-2000	data scaling
Coot	model building
PHASER	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.