5VG6

Crystal structure of D-isomer specific 2-hydroxyacid dehydrogenase from Xanthobacter autotrophicus Py2 in complex with NADPH and MES.

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	APS BEAMLINE 19-ID
Synchrotron site	APS
Beamline	19-ID
Temperature [K]	100
Detector technology	CCD
Collection date	2015-04-04
Detector	ADSC QUANTUM 315r
Wavelength(s)	0.9793
Spacegroup name	P 21 21 2
Unit cell lengths	132.514, 311.036, 98.977
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	50.000 - 2.200
R-factor	0.2013
Rwork	0.200
R-free	0.23340
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	4xa8
RMSD bond length	0.009
RMSD bond angle	1.334
Data reduction software	HKL-3000
Data scaling software	HKL-3000
Phasing software	HKL-3000
Refinement software	REFMAC

Data quality characteristics

	Overall	Inner shell	Outer shell
Low resolution limit [Å]	50.000	50.000	2.240
High resolution limit [Å]	2.200	5.970	2.200
Rmerge	0.114	0.049	0.753
Rmeas	0.128	0.055	0.850
Rpim	0.055	0.024	0.389
Number of reflections	207487		10251
<I/σ(I)>	5.1		1.9
Completeness [%]	100.0	99.8	100
Redundancy	5.3	5.1	4.7
CC(1/2)	0.800	0.997	0.801

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP	6.5	289	0.2 ul of 14 mg/ml protein in 20 mM HEPES pH 7.5, 150 mM NaCl, 10% Glycerol, 0.1% Sodium Azide, 0.5 mM TCEP, 50mM D-lactate, and 5 mM NADPH were mixed with 0.2 ul of the MCSG-2 condition #13 (0.1M MES monohydrate pH=6.5, 12%w/v PEG 20K) and equilibrated against 1.5 M NaCl solution in 96 Well 3 drop Crystallization Plate (Swissci). Before crystallization protein was incubated with 1/50 v/v of 1 mg/ml TEV solution at 289 K for 1 hour