Useful Crystallization Additives

Useful crystallization additives with relative advantages and disadvantages.

Additive

Glycerol

Advantage

Disadvantage

By increasing viscosity, it In high reduces turbulent motions in concentrations, the solution and promotes greater than 25%, the growth of fewer crystals may inhibit crystal of larger size. Cryoprotectant growth

Conditions

1–25% vol/vol of the protein solution

Reduce dielectric constant of the medium. The modification of the dielectric Alcohols and dioxane constant induces changes of poison the crystals the various interactions and inhibit (intramolecular solute– nucleation. Volatile solute, solvent–solute and so alcohols may cause Alcohols, acetone on). It has been verified 2–25% vol/vol of the problems in and dioxane experimentally that for a protein solution transferring the given concentration of crystals from the electrolyte, the reduction of solution to the ionic strength decreases goniometer head for macromolecular solubility data collection under salting-in condition and facilitates the protein crystallization Divalent cations such as Mg2+, Ca2+, Zn2+ and Cd2+ (avoid redox active cations)

Metal ions sometimes stabilize the protein structure, help the folding and may promote intermolecular interactions

Bind mainly to basic Anions such as residues. Decrease the bromide, fluoride, protein solubility, easing formate, crystallization. Bromide, malonate, iodide and anions containing acetate and heavy atoms allow citrate, single/multiwavelength phosphate and anomalous diffraction thiocyanide phasing Detergents such Are reported to promote as -octylglucoside single crystal formation, to ( -OG) improve reproducibility and crystal quality

May cause protein aggregation and precipitation

Concentration comparable with that of the protein

Excessive insolubilization of the protein might lead to amorphous precipitate

Any concentration from 5 to 200 M or even higher, depending on the anion and on the protein

Makes more difficult the handling of the solutions in which it is present (foam formation)

The initial concentration of detergent in the drop should be only slightly higher than the critical micellar concentration (CMC). It may prove advantageous to mix two detergents or add NaCl

Additive

Advantage

Disadvantage

Conditions (50–150 mM) along with the detergent. Salt concentration strongly affects the CMC of the detergent Amphiphiles such as heptanetriol act as cosurfactants and should be used for finetuning

Spermine, cadaverine, spermidine and putrescine

Counterions for negatively charged groups shield electrostatic repulsions

Ligands to metal cofactors might displace metal ions from the protein

1–50 mM