6XG4

X-ray structure of Escherichia coli dihydrofolate reductase L28R mutant in complex with trimethoprim

X-RAY DIFFRACTION

Starting Model(s)

Initial Refinement Model(s)
Type	Source	Accession Code	Details
experimental model	Other		WT DHFR solved in the same space group

Crystallization

Crystalization Experiments
ID	Method	pH	Temperature	Details
1	VAPOR DIFFUSION, HANGING DROP	5.6	293	0.1 M sodium citrate tribasic dihydrate (pH 5.6), 0.15 M ammonium acetate and 17.5% or 20% PEG 4000; 10 mM NADPH, 2 mM TMP was incubated with the L28R variant of DHFR overnight at 293 K

Crystal Properties
Matthews coefficient	Solvent content
3.03	59.39

Crystal Data

Unit Cell
Length ( ? )	Angle ( ? )
a = 61.548	�� = 90
b = 61.548	�� = 90
c = 104.661	�� = 120

Symmetry
Space Group	P 32 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 6M		2018-07-12	M	SINGLE WAVELENGTH

Radiation Source
ID #	Source	Type	Wavelength List	Synchrotron Site	Beamline
1	SYNCHROTRON	APS BEAMLINE 19-ID	0.97919	APS	19-ID

Data Collection

Overall
ID #	Resolution (High)	Resolution (Low)	Percent Possible (Observed)	Rrim I (All)	Rpim I (All)	CC (Half)	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	2.1	50	99.9	0.076	0.025	1	37.5	9.3		13880		-3

Highest Resolution Shell
ID #	Resolution (High)	Resolution (Low)	Percent Possible (All)	Percent Possible (Observed)	Rrim I (All)	Rpim I (All)	CC (Half)	Mean I Over Sigma (Observed)	Redundancy	Number Unique Reflections (All)
1	2.1	2.12	100		8.94	3.012	0.672	0.7	8.8

Refinement

Statistics
Diffraction ID	Structure Solution Method	Cross Validation method	Starting model	Resolution (High)	Resolution (Low)	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	THROUGHOUT	WT DHFR solved in the same space group	2.1	37.37	9837		70.95	0.2493	0.2493	0.2779	FULL CROSS VALIDATION	35.198

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]
-0.11	-0.05		-0.11		0.35

RMS Deviations
Key	Refinement Restraint Deviation
r_dihedral_angle_2_deg	33.498
r_dihedral_angle_3_deg	18.538
r_dihedral_angle_4_deg	13.371
r_dihedral_angle_1_deg	7.56
r_angle_refined_deg	1.417
r_angle_other_deg	1.112
r_chiral_restr	0.054
r_bond_refined_d	0.006
r_gen_planes_refined	0.006
r_bond_other_d	0.001

RMS Deviations
Key	Refinement Restraint Deviation
r_dihedral_angle_2_deg	33.498
r_dihedral_angle_3_deg	18.538
r_dihedral_angle_4_deg	13.371
r_dihedral_angle_1_deg	7.56
r_angle_refined_deg	1.417
r_angle_other_deg	1.112
r_chiral_restr	0.054
r_bond_refined_d	0.006
r_gen_planes_refined	0.006
r_bond_other_d	0.001
r_gen_planes_other	0.001

Non-Hydrogen Atoms Used in Refinement
Non-Hydrogen Atoms	Number
Protein Atoms	1271
Nucleic Acid Atoms
Solvent Atoms	131
Heterogen Atoms	76

Software

Software
Software Name	Purpose
REFMAC	refinement
PDB_EXTRACT	data extraction
HKL-3000	data reduction
HKL-3000	data scaling
MOLREP	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 R01GM157729.