7Q07

Ketol-acid reductoisomerase from Methanothermococcus thermolithotrophicus in the open state with NADP and tartrate

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	SLS BEAMLINE X10SA
Synchrotron site	SLS
Beamline	X10SA
Temperature [K]	100
Detector technology	PIXEL
Collection date	2016-06-24
Detector	DECTRIS PILATUS 6M
Wavelength(s)	0.97916
Spacegroup name	I 2 3
Unit cell lengths	130.827, 130.827, 130.827
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	41.370 - 2.200
R-factor	0.19
Rwork	0.186
R-free	0.21230
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	7q03
Data reduction software	XDS
Data scaling software	SCALA
Phasing software	PHENIX
Refinement software	PHENIX (1.19.2_4158)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	46.250	2.320
High resolution limit [Å]	2.200	2.200
Rmerge	0.163	0.528
Rmeas	0.168	0.554
Rpim	0.039	0.166
Number of reflections	19073	2766
<I/σ(I)>	23.7	4.4
Completeness [%]	100.0	100
Redundancy	18.2	10.9
CC(1/2)	0.998	0.385

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP	6.5	291.15	KARI was crystallized at 13.5 mg/ml in the following buffer 25 mM Tris/HCl pH 7.6, 2 mM dithiothreitol, 10% glycerol. The reservoir was filled with 90 ul crystallization solution (33 % (w/v) PEG 5000 MME, 100 mM MES/NaOH pH 6.5 and 200 mM ammonium sulphate). Drops of 0.7 ul protein with 0.7 ul of crystallisation solution were applied on the shelf. Crystals were firstly soaked in the crystallisation solution supplemented with 20 mM NADP, 50 mM L-Tartrate and 50 mM MgCl2 for 3 minutes, and then soaked in the crystallisation solution supplemented with 30 % v/v glycerol before freezing.