3QZU

Crystal structure of Bacillus subtilis Lipase A 7-fold mutant; the outcome of directed evolution towards thermostability

X-RAY DIFFRACTION

Crystallization

Crystalization Experiments
ID	Method	pH	Temperature	Details
1	VAPOR DIFFUSION, HANGING DROP	6	293	24% (w/v) PEG 3350, 0.2 M (NH4)2)SO4, 0.1 M BisTris-HCl, 0.03 M BES-NaOH, 0.03% (w/v) NaN3, pH 6.0, VAPOR DIFFUSION, HANGING DROP, temperature 293K

Crystal Properties
Matthews coefficient	Solvent content
2.66	53.78

Crystal Data

Unit Cell
Length ( Å )	Angle ( ˚ )
a = 59.163	α = 90
b = 45.511	β = 101.1
c = 77.613	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	CCD	MARMOSAIC 225 mm CCD	mirrors	2010-06-10	M	SINGLE WAVELENGTH

Radiation Source
ID #	Source	Type	Wavelength List	Synchrotron Site	Beamline
1	SYNCHROTRON	ESRF BEAMLINE ID23-2	0.8726	ESRF	ID23-2

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	1.71	45.51	98.3	0.159	6.8	4.1	43880	43134		3	16.9

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	1.71	1.81	88.2		0.822	1.5	3.3	18453

Refinement

Statistics
Diffraction ID	Structure Solution Method	Cross Validation method	Starting model	Resolution (High)	Resolution (Low)	Number Reflections (All)	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	PDB ENTRY 1I6W	1.85	76.16	33215	32229	1700	97.03	0.17525	0.17274	0.22275	RANDOM	13.792

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]
-0.6		0.37	0.15		0.59

RMS Deviations
Key	Refinement Restraint Deviation
r_dihedral_angle_2_deg	33.835
r_dihedral_angle_3_deg	10.42
r_dihedral_angle_4_deg	10
r_dihedral_angle_1_deg	5.851
r_scangle_it	3.238
r_scbond_it	1.967
r_mcangle_it	1.145
r_angle_refined_deg	1.107
r_mcbond_it	0.621
r_chiral_restr	0.078

RMS Deviations
Key	Refinement Restraint Deviation
r_dihedral_angle_2_deg	33.835
r_dihedral_angle_3_deg	10.42
r_dihedral_angle_4_deg	10
r_dihedral_angle_1_deg	5.851
r_scangle_it	3.238
r_scbond_it	1.967
r_mcangle_it	1.145
r_angle_refined_deg	1.107
r_mcbond_it	0.621
r_chiral_restr	0.078
r_bond_refined_d	0.011
r_gen_planes_refined	0.005

Non-Hydrogen Atoms Used in Refinement
Non-Hydrogen Atoms	Number
Protein Atoms	2700
Nucleic Acid Atoms
Solvent Atoms	482
Heterogen Atoms	42

Software

Software
Software Name	Purpose
REFMAC	refinement

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.