Crystallization 4:
Crystals of a protein were obtained by the microbatch method using
Nunc HLA plates (Nalge Nunc International). Each crystallization drop
was prepared by mixing 1.0 μl precipitant solution (12.5% PEG 4000,
0.1 M MES, pH 5.9) and 1.0μl protein solution (25 mg/ml in 20 mM Tris
HCl, 0.2 M NaCl, pH 8.0). The crystallization drop was overlaid with
a 1:1 mixture of silicone/paraffin oils, allowing slow evaporation of
the drop at 296 K.
Cryo-conditions for crystal 4:
Prior to data collection, single crystals were soaked in three
different cryoprotectant solutions for 1 min each and then transferred
to liquid nitrogen. The cryoprotectant solutions contained reservoir
solution supplemented with 5, 20, and 25% glycerol.
_exptl_crystal.crystal_id 4
_exptl_crystal.size_max ?
_exptl_crystal.size_mid ?
_exptl_crystal.size_min ?
_exptl_crystal_grow.crystal_id 4
_exptl_crystal_grow.method 'microbatch'
_exptl_crystal_grow.method_details
; Droplets were in Nunc HLA plates and covered with a 1:1 mixture of
silicone and paraffin oils, which allowed slow evaporation.
;
_exptl_crystal_grow.temp 296
loop_
_pdbx_exptl_crystal_grow_sol.crystal_id
_pdbx_exptl_crystal_grow_sol.sol_id
_pdbx_exptl_crystal_grow_sol.volume
_pdbx_exptl_crystal_grow_sol.volume_units
_pdbx_exptl_crystal_grow_sol.pH
4 'protein' 1 'microlitre' 8.0
4 'precipitant' 1 'microlitre' 5.9
loop_
_pdbx_exptl_crystal_grow_comp.crystal_id
_pdbx_exptl_crystal_grow_comp.sol_id
_pdbx_exptl_crystal_grow_comp.comp_id
_pdbx_exptl_crystal_grow_comp.comp_name
_pdbx_exptl_crystal_grow_comp.conc
_pdbx_exptl_crystal_grow_comp.conc_range
_pdbx_exptl_crystal_grow_comp.conc_units
4 'protein' 1 'protein' 25. . 'mg/ml'
4 'protein' 2 'Tris HCl' 20. . 'millimolar'
4 'protein' 3 'NaCl' 0.2 . 'molar'
4 'precipitant' 1 'PEG 4000' 12.5 . 'percent_weight_by_volume'
4 'precipitant' 2 'MES' 0.1 . 'molar'
_pdbx_exptl_crystal_cryo_treatment.crystal_id 4
_pdbx_exptl_crystal_cryo_treatment.final_solution_details
; 25% (v/v) glycerol in precipitant solution
;
_pdbx_exptl_crystal_cryo_treatment.soaking_details
; A series of 1 min soaks beginning at 5% (v/v) glycerol in precipitant,
transiting to 20% (v/v) glycerol, and finally to 25% glycerol
;
_pdbx_exptl_crystal_cryo_treatment.cooling_details
; Direct immersion in liquid nitrogen.
;
_pdbx_exptl_crystal_cryo_treatment.annealing_details
; None.
;
How this example will appear in the journal
| Crystallization |
| Crystallization method
|
Microbatch
|
| Temperature
|
296
|
| Additional details
|
Droplets were in Nunc HLA plates and covered with a
1:1 mixture of silicone and paraffin oils, which allowed
slow evaporation.
|
| Crystallization solutions |
| Protein (1 μl)
|
Protein 25 mg ml-1,
Tris 20 mM,
NaCl 0.2 M, pH 8.0
|
| Precipitant (1 μl)
|
PEG 4000 12.5% (w/v),
MES 0.1 M
pH 5.9
|
| Cryo treatment
|
| Final cryoprotection solution
|
25% (v/v) glycerol in precipitant solution
|
| Soaking
|
A series of 1 min soaks beginning at 5% (v/v)
glycerol in precipitant, transiting to 20% (v/v)
glycerol, and finally to 25% glycerol
|
| Cooling
|
Direct immersion in liquid nitrogen
|
| Annealing
|
None
|
|
|
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