Albumin isoelectric point

Human serum albumin has been purified similarly with 25% EtOH and 0.2% decanol. The isoelectric points of bovine and human serum albumins are 5.1 and 4.9. 

Some like casein, salt free globulin and acid albumin are not heavily solvated in solution and are thus readily precipitated at the isoelectric point in a manner similar to the suspensions already considered. Others, e.g. glutin, gelatine and natural albumin, are solvated like silica which at the isoelectric point are not necessarily precipitated being maintained in the sol form by the solvent. On removal of the stabilising water however by the addition of alcohol or neutral salts precipitation will occur and this most readily at the isoelectric point. 

Both isoelectric albumin and its undissociated acid and basic salts are less solvated and imbibe less liquid than the ionised form thus the viscosity of an albumin solution will be least at the isoelectric point rising to more or less well defined maxima on each side, these maxima coinciding with the points of maximum dis-... 

Albumin did not demonstrate any corrosive behavior to the copper and nickel films. The intensity of the 2100 cm 1 water association band, shown in Figure 4, did not change during the 4-hr period that albumin was initially exposed to the thin metal films. Even after 4 additional hours of exposure to the adsorbed protein film, the 2100 cm 1 band intensity remained unchanged, indicating that the copper and nickel films remained intact. Exposure of copper and nickel films to albumin above and below the protein isoelectric point did not alter these results. 

Phosphate buffers are generally used for the chromatography their initial concentration is close to 1 mM and the pH, between 6.5 and 6.9, remains constant. Elution of antibodies is obtained by phosphates, citrates, and fluorides. The most common way to desorb is, in fact, to apply a gradient of phosphate buffer at constant pH. When an antibody with a high isoelectric point is applied, the resolution power of phosphate may not be sufficient enough to separate albumin from the antibody.123... 

Theoretically, the precipitation of a protein, which is built up of amino-acids and functions as a typical amphoteric electrolyte, should best be accomplished at the isoelectric point, that is, at the hydrogen ion concentration at which the acidic and basic functions of the protein are equal and at a minimum. According to Soerensen, the iso-electric point of egg albumin is at pH 4.8,10 and while precipitation would undoubtedly be most complete at this point, it is not necessarily the most favorable for crystallization, as the egg albumin crystals are not isoelectric protein, but a compound of this with the sulfate ion, and perhaps the ammonium ion as well.11... 

Specific interactions between starch and proteins were observed as early as the beginning of the twentieth century. Berczeller996 noted that the surface tension of aqueous soap solutions did not decrease with the addition of protein (egg albumin) alone, but it did decrease when starch and protein were added. This effect was observed to increase with time. Sorption of albumin on starch is inhibited by bi- and trivalent ions and at the isoelectric point. Below the isoelectric point, bonding between starch and albumin is ionic in character, whereas nonionic interactions are expected above the isoelectric point.997 The Terayama hypothesis998 predicts the formation of protein complexes with starch, provided that starch exhibits the properties of a polyelectrolyte. Apart from chemically modified anionic starches (such as starch sulfate, starch phosphate, and various cross-linked starch derivatives bearing ionized functions), potato starch is the only variety that behaves as a polyelectrolyte. Its random phosphate ester moieties permit proteins to form complexes with it. Takeuchi et a/.999-1002 demonstrated such a possibility with various proteins and a 4% gel of potato starch. 

At the second critical pH (pH,, ), which is usually below the protein isoelectric point, strong electrostatic interaction between positively charged protein molecules and anionic polysaccharide chains will cause soluble protein/polysaccharide complexes to aggregate into insoluble protein/polysaccharide complexes. For negatively charged weak acid-based (e.g., carboxylic acid) polysaccharides like pectin, with the decrease of pH below the pKa of the polysaccharide, protein (e.g., bovine serum albumin (BSA))/polysaccharide (e.g., pectin) insoluble complexes may dissociate into soluble complexes, or even non-interacted protein molecules and polysaccharide chains, due to the low charges of polysaccharide chains as well as the repulsion between the positively charged proteins (Dickinson 1998). 

In a related connection, the solubility of a protein may be considerably greater in a slightly acidified or alkalized weakly protic solvent than in the pure liquid. This would be analogous to the situation which one obtains in aqueous media, in which proteins are generally increasingly soluble the further the pH is from the isoelectric point of the protein, within certain pH limits (Cohn and Edsall, 1943, p. 606). This is probably the reason that 2-chloroethanol is such an excellent solvent for proteins (Doty, 1959). This solvent is not a very stable one, and significant amounts of HCl can be present in it. This may also account for the observation that, although bovine serum albumin is insoluble in pure acetone, methanol, and ethanol, it dissolves in them when trichloroacetic acid (1%) is added (Levine, 1954). On the other hand, the trichloroacetate counterion may itself influence the solubility of the protein salt in the nonaqueous solvents. 

Results of adsorption experiments are presented in Table 2. In solution, TNF-a, a polypeptide with MW 17 kD, exists as a trimer with MW 51 kD. The monomeric units are kept together due to the hydrophobic interaction. From the experimental data it is clear that large mesopores play a relatively minor role in adsorption of TNF-a, as both MN resins adsorb it rather efficiently. Isoelectric point of TNFa is similar to that of serum albumin (pi between 4.8 and 5.6) [11], It has been shown that TNFa can be removed from biological media using non-polar cross-linked polystyrene adsorbents of the Amberlite type (XAD-2, XAD-4 and XAD-6) [12], This observation confirms that hydrophobic interactions play an important role in the adsorption of TNFa at neutral pH. Our data support this conclusion. 

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