Immunochemical studies on thermal denaturation of ovomucoid.
The thermal denaturation of ovomucoid was investigated by immunochemical methods, namely immunoprecipitation analyses and antibody-Sepharose 4B column chromatography. In the immunoprecipitation analyses, heated ovomucoid (90 degrees C, 90 min, pH 7.2) required about twice the antigen addition of the native protein to approach maximal precipitation with specific antibody, and the maximal immunoprecipitation was decreased to 80% of that by native ovomucoid. However, heated protein inhibited the binding of antibody with native ovomucoid, and 100% inhibition was attained at about 4-times the antigen addition necessary for the native protein. Heated ovomucoid (100 degrees C, 120 min) showed little immunoprecipitation and inhibition. To ovomucoid antigenicity was diminished more slowly than the trypsin inhibitory activity by heating, e.g., heated ovomucoid (90 degrees C, 120 min) retained more than 30% of the antigenicity but little trypsin inhibitory activity. By passing through the immunoaffinity column, heated ovomucoid (90 degrees C, 90 min) was separated into two fractions, either with (fraction II) or without (fraction I) antigenicity. Fraction II contained smaller fractions of ordered secondary structure than native ovomucoid, and trypsin inhibitory activity of fraction II was only 24% of the native one. These results indicated that thermally denatured ovomucoid was heterogeneous regarding the conformational damage caused by heating, and the structure around some antigenic sites in an ovomucoid molecule was retained even after the backbone conformation was partially destroyed and trypsin inhibitory activity was lost.
D000818:Animals; D000906:Antibodies; D000936:Antigen-Antibody Complex; D002645:Chickens; D002942:Circular Dichroism; D004527:Egg Proteins / Q000276:immunology; D004529:Egg White; D000939:Epitopes / Q000032:analysis; D005260:Female; D006358:Hot Temperature; D005779:Immunodiffusion; D010059:Ovomucin / Q000276:immunology / Q000494:pharmacology; D011487:Protein Conformation; D011489:Protein Denaturation; D014357:Trypsin / Q000378:metabolism
D000906:Antibodies; D000936:Antigen-Antibody Complex; D004527:Egg Proteins; D000939:Epitopes; D010059:Ovomucin; D014357:Trypsin