Ovomucoid denaturation

Egg proteins contain many fractions, of which ovoalbumin, ovomucoid, and livetin (an egg-yolk allergen) are the strongest allergens. They are resistant to digestion, thermal denaturation and low pH (for example the allergic properties of ovoalbumin can still be observed at pH 3.0). 

These proteins are composed of simple proteins in combination with carbohydrates like mucopolysaccharides, which include hyaluronic acid and chondroitin sulphates. On hydrolysis, mucopolysaccharides yield more than 4% of amino-sugars, hexosamine and uronic acid e.g., ovomucoid from egg white. Soluble mucoproteins are neither readily denatured by heat nor easily precipitated by common protein precipitants like trichloroacetic acid or picric acid. The term glycoproteins is restricted to those proteins that contain small amounts of carbohydrate usually less than 4% hexosamine. 

HiroseJ, KitabatakeN, KimuraA, NaritaH (2004). Recognition of native and/or thermally induced denatured forms of the major food allergen, ovomucoid, by human IgE and mouse monoclonal IgG antibodies. Biosci. Biotechnol Biochem., 68 2490-2497. 

Pspsin (EC 3.4.23.1) a protease in the stomach of all vertebrates with the exception of stomachless fish (e.g. carp). Purified P. shows maximal activity at pH 1-2, but in the stomach the optimal pH is 2-4. Above pH 6, P. is inactivated by denaturation. It preferentially catalyses hydrolysis of peptide bonds between two hydrophobic amino acids (Phe-Leu, Phe-Phe, Phe-TyrT With the exception of protamines, keratin, mucin, ovomucoid and other carbohydrate-rich proteins, most proteins are attacked by P. The products of P. action are peptone, i. e. mixtures of peptides in the M range 300-3,000. P. is a highly acidic (pi 1), single chain phosphoprotein (327 amino acid residues of known primary sequence, M, 34,500), which is released from its zymogen (pepsinogen, 42,500) by autocatalysis in the presence of hydrochloric acid. 

Many enzymes or other proteins are stabilized by interaction with small ligands (substrates, cofactors, inhibitors, products). Nonspecific protection by macromolecules and stabilization by specific protein—protein interaction are generally less readily quantified. As a model system in which to determine effects of protein—protein interaction on conformational (thermal) stability, Donovan and Beardslee (1975) selected the proteinase—inhibitor association between bovine 3-trypsin and STI or chicken ovomucoid. They observed that both protein components were stabilized in the 1 1 complexes and, furthermore, that each trypsin—inhibitor complex showed one endotherm in the DSC. No stabilization was observed for a weaker electrostatic complex between ovalbumin andjysozyme. They interpreted the stabilization as an increase in kinetic thermal stability, i.e., a decrease in the rate of denaturation at a given temperature. Results obtained by other methods (Edelhoch and Steiner, 1965 Laskowski and Sealock, 1971 Jibson et al., 1981) show that proteinase—inhibitor association generally involves no major conformational changes in the proteins. 

In egg white, however, stabilities of ovomucoid and ovoinhibitor may differ. When Matsuda et al. (1981c) heated egg white and determined the extent of aggregation by gel electrophoresis, they found that ovomucoid bands did not change after heating at 76 C and pH 7, but that ovoinhibitor bands diminished after heating at 62°C. This apparent discrepancy might be explained if ovoinhibitor interacts more readily than ovomucoid with other, readily denatured constituents of egg white to form aggregates. Hence the difference may not reflect irreversible denaturation of ovoinhibitor as such. 

Ovomucoid and ovomucin are the proteins responsible for the viscosity and gel-like consistency of egg white. Ovomucoid (28 kDa, p7=4.4-4.6) is composed of three components which differ in carbohydrate composition on the JV-terminus of the molecule. Ovomucoid inhibits trypsin and other proteolytic enzymes and is a strong allergen. The stability to thermal denaturation (in an acidic medium) is associated with nine disulfide bonds in the molecule. 

Native ovoflavoprotein (49 kDa, pf=5.1) has, as does ovomucoid, certain antinutritional effects, as it inhibits serine proteases (trypsin, chymotrypsin and also microbial proteases) and has antiviral activity. Ovomacroglobulin (ovostatin) is an inhibitor of serine, cysteine, thiol and metalloproteases and shows antimicrobial activity. Some antinutritional effects are also seen in the basic glycoprotein avidin in raw egg white (relative molecular weight of the monomer is 15.6 kDa). It contains four identical subunits (pf = 9.5), each of which binds one molecule of biotin to give an unavailable complex. However, the denatured avidin, for example in hard-boiled eggs, does not interact with biotin. The interaction of riboflavin with flavoprotein (32 kDa, pf = 4.0) has, on the contrary, a positive influence on vitamin stability. Cystatin acts as cysteine protease inhibitor, and shows antimicrobial, antitumor and immunomodulating activities. 

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