3I80

Protein Tyrosine Phosphatase 1B - Transition state analog for the second catalytic step

X-RAY DIFFRACTION

Crystallization

Crystalization Experiments
ID	Method	pH	Temperature	Details
1	VAPOR DIFFUSION, SITTING DROP	7.5	277	Drop: 2 uL of protein solution, 0.5 uL sucrose 30% (w/v) and 3 uL of precipitant solution (0.1 M HEPES pH 7.5, 0.2 M magnesium acetate and 18-20% polyethylene glycol 8000). Well: 500 uL of precipitant solution. The protein solution was prepared as follows: 9 uL of 50 mM of Na3VO4 and 1 uL of 50 mM of DADEYL peptide (at pH 8.5-9.0) were mixed and allowed to react for 1-1.5 hour; then, 50 uL of native PTP1B (12 mg/mL in 10 mM Tris pH 7.5, 25 mM NaCl, 0.2 mM EDTA and 3 mM DTT) was added and the solution used immediately for crystallization. VAPOR DIFFUSION, SITTING DROP, temperature 277K

Crystal Properties
Matthews coefficient	Solvent content
3.09	60.24

Crystal Data

Unit Cell
Length ( Å )	Angle ( ˚ )
a = 87.84	α = 90
b = 87.84	β = 90
c = 103.79	γ = 120

Symmetry
Space Group	P 31 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	IMAGE PLATE	RIGAKU RAXIS IV	mirrors	2008-09-06		SINGLE WAVELENGTH

Radiation Source
ID #	Source	Type	Wavelength List	Synchrotron Site	Beamline
1	ROTATING ANODE	RIGAKU RU200	1.5418

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	2.25	38.04	100	0.084	8.6	4.01	22453	22453			44.7

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)
	2.25	2.33	99.9		0.473	2.2	4.04	2199

Refinement

Statistics
Diffraction ID	Structure Solution Method	Starting model	Resolution (High)	Resolution (Low)	Cut-off Sigma (F)	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	PDB ENTRY 1SUG	2.25	35.713	1.9	22441	22441	1149	52.79	0.201	0.201	0.199	0.235	RANDOM	45.377

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]
-0.669			-0.669		1.339

RMS Deviations
Key	Refinement Restraint Deviation
f_dihedral_angle_d	17.173
f_angle_d	0.939
f_chiral_restr	0.067
f_bond_d	0.006
f_plane_restr	0.004

Non-Hydrogen Atoms Used in Refinement
Non-Hydrogen Atoms	Number
Protein Atoms	2434
Nucleic Acid Atoms
Solvent Atoms	231
Heterogen Atoms	31

Software

Software
Software Name	Purpose
d*TREK	data scaling
PHASER	phasing
PHENIX	refinement
PDB_EXTRACT	data extraction
CrystalClear	data collection
d*TREK	data reduction

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.