4YQ3

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-F
Synchrotron site	APS
Beamline	21-ID-F
Temperature [K]	100
Detector technology	CCD
Collection date	2010-11-04
Detector	MARMOSAIC 225 mm CCD
Wavelength(s)	0.97872
Spacegroup name	H 3 2
Unit cell lengths	98.062, 98.062, 176.131
Unit cell angles	90.00, 90.00, 120.00

Refinement procedure

Resolution	41.279 - 2.490
R-factor	0.1813
Rwork	0.177
R-free	0.21930
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	1p9p
RMSD bond length	0.003
RMSD bond angle	0.706
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	2.180
High resolution limit [Å]	2.100	4.520	2.100
Rmerge	0.146	0.049	0.591
Total number of observations	126326
Number of reflections	11684
<I/σ(I)>	7.8
Completeness [%]	99.5	99.5	97.6
Redundancy	6.5	7	3.9

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.