1.4 Proteins 61polar solvents such as water, glycerol, for-
mamide, dimethylformamide or formic acid.
In a less polar solvent such as ethanol, pro-
teins are rarely noticeably soluble (e. g.
prolamines). The solubility in water is de-
pendent on pH and on salt concentration.
Figure 1.36 shows these relationships for
β-lactoglobulin.
At low ionic strengths, the solubility rises with
increase in ionic strength and the solubility mini-
mum (isoelectric point) is shifted from pH 5.4to
pH 5.2. This shift is due to preferential binding of
anions to the protein.
If a protein has enough exposed hydrophobic
groups at the isoelectric point, it aggregates
due to the lack of electrostatic repulsion
via intermolecular hydrophobic bonds, and
(isoelectric) precipitation will occur. If on
the other hand, intermolecular hydropho-
bic interactions are only poorly developed,
a protein will remain in solution even at the
isoelectric point, due to hydration and steric
repulsion.
As a rule, neutral salts have a two-fold effect on
protein solubility. At low concentrations they in-
crease the solubility (“salting in” effect) by sup-
pressing the electrostatic protein-protein interac-
tion (binding forces).
Fig. 1.36.β-Lactoglobulin solubility as affected by pH
and ionic strength I. 0.001, II. 0.005, III. 0.01, IV. 0. 02
The log of the solubility (S) is proportional
to the ionic strength (μ) at low concentrations
(cf. Fig. 1.36.):logS=k·μ. (1.95)Protein solubility is decreased (“salting out” ef-
fect) at higher salt concentrations due to the ion
hydration tendency of the salts. The following re-
lationship applies (S 0 : solubility at μ=0; K: salt-
ing out constant):logS=logS 0 −K·μ. (1.96)Cations and anions in the presence of the same
counter ion can be arranged in the following or-
ders (Hofmeisterseries) based on their salting out
effects:K+>Rb+>Na+>Cs+>Li+>NH+ 4 ;SO 42 −>citrate^2 −>tratrate^2 −>acetate−
>CI−>NO− 3 >Br−>J−>CNS−. (1.97)Multivalent anions are more effective than mono-
valent anions, while divalent cations are less ef-
fective than monovalent cations.
Since proteins are polar substances, they are
hydrated in water. The degree of hydration
(g water of hydration/g protein) is variable.
It is 0.22 for ovalbumin (in ammonium sul-
fate), 0.06 for edestin (in ammonium sulfate),
0 .8forβ-lactoglobulin and 0.3 for hemoglobin.
Approximately 300 water molecules are suffi-
cient to cover the surface of lysozyme (about
6000 Å^2 ), that is one water molecule per
20 Å^2.
The swelling of insoluble proteins corresponds
to the hydration of soluble proteins in that
insertion of water between the peptide chains
results in an increase in volume and other
changes in the physical properties of the pro-
tein. For example, the diameter of myofibrils
(cf. 12.2.1) increases to 2.5 times the original
value during rinsing with 1.0mol/LNaCl,
which corresponds to a six-fold volume in-
crease (cf. 12.5). The amount of water taken
up by swelling can amount to a multiple of
the protein dry weight. For example, muscle
tissue contains 3.5–3.6 g water per g protein dry
matter.