8QMR

Succinic semialdehyde dehydrogenase from E. coli with bound NAD+ and succinic semialdehyde

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	ESRF BEAMLINE MASSIF-3
Synchrotron site	ESRF
Beamline	MASSIF-3
Temperature [K]	100
Detector technology	PIXEL
Collection date	2023-05-18
Detector	DECTRIS EIGER X 4M
Wavelength(s)	0.9677
Spacegroup name	P 21 21 21
Unit cell lengths	91.520, 115.620, 179.640
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	38.070 - 2.300
R-factor	0.197
Rwork	0.196
R-free	0.22780
Structure solution method	MOLECULAR REPLACEMENT
RMSD bond length	0.003
RMSD bond angle	0.540
Data reduction software	XDS
Data scaling software	XSCALE
Phasing software	PHENIX
Refinement software	PHENIX ((1.20.1_4487: ???))

Data quality characteristics

	Overall	Inner shell	Outer shell
Low resolution limit [Å]	38.070	38.070	2.360
High resolution limit [Å]	2.300	10.290	2.300
Rmerge	0.154	0.051	0.998
Rmeas	0.164	0.055	1.063
Number of reflections	85212	1051	6232
<I/σ(I)>	9.59
Completeness [%]	99.9
Redundancy	8.4
CC(1/2)	0.998	0.999	0.859

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP		295	7.3 mg/mL of SAD in 20 mM HEPES-KOH, 50 mM KCl, pH 7.5 was pre-mixed with NAD+ (final concentration 5 mM). The pre-mixed protein was then mixed in a 2:1 ratio with 250 mM Bis-Tris-Propane, pH 8.25, 20 % (w/v) PEG4000. The final drop size was 3 uL. The crystal was soaked with 5 mM succinic semialdehyde for 2 minutes and the drop was complemented with PEG200 to a final concentration of 20 % (v/v) before flash freezing the crystal in liquid nitrogen.