2W3A

HUMAN DIHYDROFOLATE REDUCTASE COMPLEXED WITH NADPH AND TRIMETHOPRIM

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	CHESS BEAMLINE A1
Synchrotron site	CHESS
Beamline	A1
Temperature [K]	100
Detector technology	CCD
Collection date	1997-07-14
Detector	ADSC QUANTUM-1
Spacegroup name	C 2 2 21
Unit cell lengths	88.468, 94.014, 96.576
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	9.900 - 1.500
R-factor	0.181
Rwork	0.179
R-free	0.21400
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	UNPUBLISHED HUMAN DHFR FOLATE COMPLEX
RMSD bond length	0.012
RMSD bond angle	1.506
Data reduction software	MOSFLM
Data scaling software	SCALA
Phasing software	AMoRE
Refinement software	REFMAC (5.2.0019)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	20.000	1.540
High resolution limit [Å]	1.500	1.500
Rmerge	0.050	0.190
Number of reflections	55140
<I/σ(I)>	8.3	2.9
Completeness [%]	85.7	28.3
Redundancy	3.3	1.1

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, HANGING DROP	8.75	277	HUMAN DHFR/FOLATE COMPLEX WAS MIXED WITH NADPH AND TRIMETHOPRIM (BOTH 2 MM FINAL). CRYSTALS WERE GROWN BY HANGING DROP VAPOR DIFFUSION AT 277 K BY MIXING EQUAL VOLUMES OF PROTEIN/NADPH/TMP WITH RESERVOIR (24% PEG 4000, 200 MM LI2SO4, 100 MM TRIS.HCL, PH 8.75). TRUNCATED TRIANGULAR CRYSTALS APPEARED SLOWLY, IN ABOUT A MONTH. THE CRYSTAL WAS CRYOPROTECTED WITH 15% GLYCEROL AND FLASH-COOLED IN LIQUID N2.