Ionic strength reducers: an efficient approach to protein purification and crystallization. Application to two Rop variants.
Detailed knowledge of the influence of various parameters on macromolecular solubility is essential for crystallization. The concept of so-called 'ionic strength reducers' provides insight into the changes in solubility induced by organic solvents and hydrophilic polymers in aqueous electrolytic solutions. A simple and efficient procedure is presented which exploits the properties of ionic strength reducers in the purification and crystallization of proteins. Using two designed variants of the Rop protein as model systems, superior crystals have been obtained compared with conventional techniques. This procedure is particularly useful in cases where excessive nucleation leads to the growth of a large number of tiny crystals that are useless for crystallographic analysis.
Papanikolau
Yannis
Y
University of Crete, Department of Biology, PO Box 2208, GR-71409 Heraklion, Crete, Greece.
Kotsifaki
Dina
D
Fadouloglou
Vasiliki E
VE
Gazi
Anastasia D
AD
Glykos
Nicholas M
NM
Cesareni
Giovanni
G
Kokkinidis
Michael
M
D000409:Alanine / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D003460:Crystallization; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics; D009154:Mutation / Q000235:genetics*; D009994:Osmolar Concentration; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*
D001426:Bacterial Proteins; D016601:RNA-Binding Proteins; C054243:Rop protein, ColE1 plasmid; D000409:Alanine