4YX7

Complex of SpaO(SPOA1,2) and OrgB(APAR)::T4lysozyme fusion protein

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	NSLS BEAMLINE X29A
Synchrotron site	NSLS
Beamline	X29A
Temperature [K]	100
Detector technology	CCD
Collection date	2013-11-22
Detector	ADSC QUANTUM 315r
Wavelength(s)	1.075
Spacegroup name	P 1 21 1
Unit cell lengths	62.092, 89.070, 62.092
Unit cell angles	90.00, 114.94, 90.00

Refinement procedure

Resolution	47.590 - 2.001
R-factor	0.1586
Rwork	0.157
R-free	0.20960
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	4yx5 2lzm
RMSD bond length	0.008
RMSD bond angle	1.159
Data scaling software	Aimless (0.2.7)
Phasing software	PHENIX
Refinement software	PHENIX

Data quality characteristics

	Overall	Inner shell	Outer shell
Low resolution limit [Å]	47.590	47.590	2.050
High resolution limit [Å]	2.000	8.940	2.000
Rmerge	0.102	0.062	0.530
Rpim	0.050	0.030	0.253
Total number of observations	210953	2212	15750
Number of reflections	41204
<I/σ(I)>	10.5	19.6	3.2
Completeness [%]	99.5	92.8	99.8
Redundancy	5.1	4.9	5.2
CC(1/2)	0.994	0.991	0.828

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, HANGING DROP		293	SpaO(145-213) + SpaO (232-297) + OrgB(1-30)::T4 lysozyme was concentrated to 18.5mg/mL and crystallized with 25% PEG3350, 200mM ammonium formate, 100mM sodium acetate pH=5.0. Microseeding was employed to enhance crystal uniformity and diffraction. Briefly, crystals to be seeded were harvested in precipitant solution and vortexed in a microfuge tube with a small stir bar for ~60 seconds. The slurry of microseeds was serially dilluted (5-10-fold steps) in precipitant solution and 5 selected microseed-precipitant mixtures were mixed with fresh protein as in a normal hanging drop experiment. Crystals were cryoprotected in 30% PEG3350, 10% glycerol, 200mM ammonium acetate, 100mM sodium acetate pH=5.0.