Ovalbumin hybrid

The enzymatic synthesis of a hybrid type oligosaccharide on the hen ovalbumin with 13C-enriched galactose allowed the measurement of the 13C-NMR spectra of the whole glycoprotein and additionally, the determination of the correlation times of the protein and the oligosaccharide to 25 ns and 40-80 ns, respectively [168]. This implies that the carbohydrate, at least in its terminal monosaccharide constituents, has a much higher flexibility than the protein, and its flexibility is only little impeded by the attachment to the protein. 

Fig. 1. Estimation of the size of mRNA for bovine poly(ADP-ribose) synthetase by in vitro translation of size fractionated poly (A) + RNA and Northern blot analysis. A. Size-fractionation of poly (A) + RNA and location of mRNA for the enzyme. Bovine thymus poly(A)+ RNA was size-fractionated by neutral sucrose density gradient centrifugation and RNA in each fraction was translated in vitro. The translated products were immunoprecipitated, and separated on a 7.5% SDS-polyacrylamide gel. B. Typical fluorogram of the gel. Lanes 1, 2, 3, 4, and 5, correspond to fractions 1, 3, 5, 7, and 9, respectively. Molecular weight markers a, p-galactosidase (116K), b, phosphorylase a(95K), c, bovine serum albumin (68K), d, ovalbumin (43K), e, lysozyme (14.3K). C. Northern blot analysis of bovine thymus poly(A)+ RNA. RNA (2 pg per lane) was separated on a 1.2% agarose/formaldehyde gel, transferred to a nitrocellulose filter, and hybridized with the p-iabelled 2.7 kb insert cDNA prepared from the clone ARS-1. Size markers used were 28 S(4.9 kb), 18 S(2.0 kb)rRNA, 3.8 kb, 2.7 kb, and 1.1 kb denatured DNA. Reprinted from Taniguchi etaL, Eur J Biochem 171 571-575,1988.

Univalent, hybrid molecules of antiovalbumin and nonspecific IgG failed to fix guinea pig complement in the presence of ovalbumin, indicating that bivalence is essential (58). Although univalent fragments produced by papain also do not fix complement, this conceivably could be due to the absence of fragment Fc this alternative explanation would not apply in the case of 7 S univalent molecules. 

The initial evidence for the existence of hybrid molecules was the capacity of such preparations to precipitate a mixture of the two antigens (ovalbumin and bovine IgG) but neither individual antigen alone (89). A hybrid molecule would not be expected to form a continuous lattice with either antigen but should do so if both are present (see Fig. 6.4). The failure of molecules A-A or B-B in the mixture to precipitate the corresponding antigens was attributed to the blocking activity of A-B, which is univalent with respect to either antigen and is presumably present in twofold excess. 

Fig. 8.3. Agglutination reactions of mixtures of chicken erythrocytes (oval and nucleated) and human erythrocytes (disc-shaped) the chicken cells were eoated with bovine IgG (BGG) and the human cells with hen ovalbumin. The following rabbit F(ab >2 fragments were present in the mixtures A, antiovalbumin B, anti-BGG C, mixture of antiovalbumin and anti-BGG D, hybrid preparation of antiovalbumin and anti-BGG. From reference 94.

Figure 9.6. Structures of the third family of iV-linked oligosaccharides—hybrids of the high mannose and iV-acetyl-lactosamine families, observed in ovalbumin oligosaccharides.

It has been shown that, during secondary stimulation with estrogen, there is an increase in the concentration of nuclear receptor molecules followed by an increase in the available initiation sites for RNA synthesis on the chromatin (Tsai et al., 1975b Kalimi et al., 1976 Anderson et al., 1972). Using hybridization techniques, it has been shown that control of the expression of the ovalbumin gene by estrogen occurs at the transcriptional level (Harris et al., 1976). 

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