The solubility of a peptide in water cannot be predicted by studying its structure. However, the ε-amino group of Lys and the guanidine of Arg are usually helpful for estimating the solubility of peptides, particularly those with short sequences. In contrast, acidic peptides that contain Asp and Glu tend to be insoluble in water but can be dissolved easily in diluted ammonia or basic buffers.
Certain basic characteristics can be used to predict solubility:
Peptides containing <5 amino acids are commonly soluble in aqueous solutions. However, if the entire sequence consists of hydrophobic residues it will have only limited solubility or could be completely insoluble.
Hydrophilic peptides that contain >25% charged amino acids (E, D, K, R, and H) and <25% hydrophobic residues are usually soluble in aqueous solutions.
Hydrophobic peptides whose sequence contains ≥50% hydrophobic residues might be completely or only partially soluble in aqueous solutions. These peptides should instead be dissolved in organic solvents such as DMSO if they do not contain C, W, or M residues. If they do contain these amino acids, they should be dissolved in DMF, acetonitrile, isopropyl alcohol, ethanol, acetic acid, 4–8 M guanidine hydrochloride (GdnHCl), or urea before being diluted carefully in an aqueous solution.
Hydrophobic peptides that include >75% hydrophobic amino acids are generally not soluble in aqueous solutions. Instead, very strong solvents such as TFA or formic acid must be used for the initial solubilization. However, the peptide might precipitate when added to an aqueous buffered solution. As such, high concentrations of organic solvents or denaturants might be needed to dissolve these peptides.
Peptides that include a very high proportion (>75%) of D, E, H, K, N, Q, R, S, T, or Y can form intermolecular hydrogen bonds (cross-links), which can result in gel formation in concentrated aqueous solutions. Therefore, peptides should be dissolved in an organic solvent that is compatible with the final experiment. After dissolving the peptides in the organic solvent, the solution should be added slowly (dropwise) to a stirring aqueous buffered solution. The limit of solubility is reached when the resulting peptide solution begins to show turbidity.