4HLC

Sulfonylpiperidines as Novel, Antibacterial Inhibitors of Gram-Positive Thymidylate Kinase (TMK): Compound 5

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	APS BEAMLINE 31-ID
Synchrotron site	APS
Beamline	31-ID
Temperature [K]	100
Detector technology	CCD
Collection date	2009-10-23
Detector	RAYONIX MX225HE
Wavelength(s)	0.98
Spacegroup name	P 1 21 1
Unit cell lengths	46.200, 90.970, 48.330
Unit cell angles	90.00, 102.85, 90.00

Refinement procedure

Resolution	41.840 - 1.550
R-factor	0.1799
Rwork	0.179
R-free	0.19850
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	in-house apo-state coordinates
RMSD bond length	0.010
RMSD bond angle	1.100
Data reduction software	XDS
Data scaling software	SCALA
Phasing software	AMoRE
Refinement software	BUSTER (2.11.2)

Data quality characteristics

	Overall	Inner shell	Outer shell
Low resolution limit [Å]	42.000	42.000	1.640
High resolution limit [Å]	1.550	4.910	1.550
Rmerge	0.054	0.021	0.371
Number of reflections	55957
<I/σ(I)>	14.3	28.8	2.3
Completeness [%]	99.9	99.2	99.9
Redundancy	3.6	3.7	3.5

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP	6.8	293	To obtain the inhibitor bound crystal form of TMK-S.aureus crystals were initially grown in the absence of compound using the sitting drop method at 293 K with a reservoir solution of 100 mM PCPT (propionate-cacodylate-bistris propane buffer) pH 7-8, 21-24% PEG 3350, 200 mM Mg2Cl using 1:1 protein:reservoir solution with the protein solution at 13 mg/mL. Crystals were harvested and soaked overnight in a solution containing 100 mM PCPT, 35% PEG 3350, 200 mM Mg2Cl and 1-2 mM TK-666 from a 100 mM DMSO stock. After soaking the crystals were cryoprotected by soaking for 15 minutes in compound-soak solution supplemented with 20% ethylene glycol. , VAPOR DIFFUSION, SITTING DROP