3BOM

Crystal structure of trout hemoglobin at 1.35 Angstrom resolution

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	APS BEAMLINE 23-ID-D
Synchrotron site	APS
Beamline	23-ID-D
Temperature [K]	100
Detector technology	CCD
Collection date	2006-12-16
Detector	MARMOSAIC 300 mm CCD
Wavelength(s)	0.97935
Spacegroup name	P 1 21 1
Unit cell lengths	57.455, 63.162, 78.702
Unit cell angles	90.00, 93.10, 90.00

Refinement procedure

Resolution	39.294 - 1.350
R-factor	0.172
Rwork	0.170
R-free	0.21200
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	1ouu
RMSD bond length	0.009
RMSD bond angle	1.197
Data reduction software	DENZO
Data scaling software	SCALEPACK
Phasing software	MOLREP
Refinement software	REFMAC (5.2.0005)

Data quality characteristics

	Overall	Inner shell	Outer shell
Low resolution limit [Å]	39.294	39.294	1.380
High resolution limit [Å]	1.350	3.330	1.350
Rmerge	0.118	0.088	0.459
Number of reflections	120196
<I/σ(I)>	10.482		2.306
Completeness [%]	97.4	99	77.8
Redundancy	7	7.3	4.2

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, HANGING DROP	5.7	277	Protein solution (15 mg/mL CO-bound trout IV Hemoglobin, 0.025 M Sodium chloride, 0.010 M Tris-HCl pH 8.0) was mixed one to one with carbon monoxide flushed well solution to yield a final concentration of 22.5% PEG 1500 and 0.04-0.06 M MES/Acetate at pH 5.7. The solutions were pH-ed to verify that the crystallization conditions were at pH 5.7 due to the lower buffer molarity. Crystals were cryo-protected with 22.5% PEG 1500, 22.5% Ethylene glycol, 0.04-0.06 M MES/Acetate at pH 5.7 in a single step, VAPOR DIFFUSION, HANGING DROP, temperature 277K