4YQ9

Crystal structure of TrmD, a M1G37 tRNA Methyltransferase with SAM-competitive compounds

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	APS BEAMLINE 21-ID-G
Synchrotron site	APS
Beamline	21-ID-G
Temperature [K]	100
Detector technology	CCD
Collection date	2010-03-31
Detector	MARMOSAIC 300 mm CCD
Wavelength(s)	0.97856
Spacegroup name	H 3 2
Unit cell lengths	94.738, 94.738, 177.726
Unit cell angles	90.00, 90.00, 120.00

Refinement procedure

Resolution	37.246 - 1.640
R-factor	0.165
Rwork	0.164
R-free	0.18760
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	1p9p
RMSD bond length	0.012
RMSD bond angle	1.340
Data scaling software	HKL-2000
Phasing software	PHASER
Refinement software	PHENIX

Data quality characteristics

	Overall	Inner shell	Outer shell
Low resolution limit [Å]	50.000	50.000	1.640
High resolution limit [Å]	1.580	3.400	1.580
Rmerge	0.050	0.025	0.558
Total number of observations	259930
Number of reflections	42204
<I/σ(I)>	12.9
Completeness [%]	99.7	98.7	99.6
Redundancy	6.2	5.9	6.2

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP	7.5	295	Protein solution:( 12/mg/mL in 100mM HEPES pH 7.5, 150mM NaCl, 10mM MgCl2 2mM DTT) Well solution: (20% PEG3,350 and 0.2M potassium citrate tribasic monohydrate). 4uL of S-adenosyl methionine in water were added to 100uL of protein and allowed to incubate on ice for 1 hour before protein was mixed with well at 1:1 ratio.Seeding used to improve crystals. Compound stock solutions (either 100mM or 1M stocks) were added up to a final drop concentration of 4.8% DMSO. Crystals were soaked for 4-6 hours. 20% glycerol in well solution was used as cryoprotectant for a quick dip of crystal in liquid N2.