Glycoprotein ovalbumin

Table I summarizes the sequencing results from alcohol dehydrogenase and the N-terminally blocked glycoprotein ovalbumin. The probable identity of these fragments is indicated. All fragments identified for both proteins by N-terminal sequencing corresponded to cleavage after cystine. The data from ovalbumin are particularly interesting. The structure of ovalbumin is well characterized (5) and contains only one disulfide bond between Cys 73 and Cys 120 yet sequence was obtained following Cys 11 and Cys 30. The bands for these fragments appeared more slowly than the odiers and were fainter in appearance.

Ovalbumin constitutes about 54% of the protein in egg white. The remainder consists of lysozyme, avidin and about a dozen different glycoproteins. Ovalbumin is a phosphoprotein of molecular weight about 45,000 which contains about 0.1% P in the form of phosphate groups bound to serine units (Figure 10.28) [34],... 

A DSC study of tetrameric Con A (pH 7) by Borrebaeck and Mattiasson (1980) showed that at 2.5 metal ions per monomer. Con A was stabilized by Mn +, Ca , or both. Additional stabilization by methyl a-D-mannoside was observed primarily when both metals were present, but some was observed with Ca2+. Stabilization of Con A by small saccharides was a function of the association constant. Association with horseradish peroxidase, a glycoprotein, stabilized Con A at pH 7. In contrast, we found no stabilization on association with the egg-white glycoproteins, ovalbumin or ovomucoid, at pH 5. 

Proteins. A chiral stationary phase with immobilized a -acid glycoprotein on silica beads was introduced by Hermansson in 1983 [18, 19]. Several other proteins such as chicken egg albumin (ovalbumin), human serum albumin, and cellohy-drolase were also used later for the preparation of commercial CSPs. Their selectivity is believed to occur as a result of excess of dispersive forces acting on the more retained enantiomer [17]. These separation media often exhibit only modest loading capacity. 

The resolution of these columns for protein mixtures, however, was comparably poor. The peak capacity for human serum albumin was near 3 during 20 min gradient elution. Improvement has been reached by covalent binding of PEI (M = 400-600) onto a 330 A silica of 5 pm particle size [38], The peak capacities of ovalbumin and 2a -arid glycoprotein were 30-40 (tgradienl = 20 min). Enhanced peak capacity and resolution probably were due to the more diffuse structure of PEI coupled to silane moieties than that of strictly adsorbed on silica and cross-linked (see Sect, 2.2). Other applications of covalently adsorbed PEI are discussed in Sect. 4.1. 

Secretion of nonglycosylated macromolecules in the presence of tunicamycin has also been investigated in a number of other cells. Rat-liver cell-secretion ol albumin (a carbohydrate-free protein), transferrin, and a-acid glycoprotein was not inhibited, and, in chick-liver cells, only a decrease by 10-25% in the secretion of transferrin and the apoprotein B chain of very-low-density lipoprotein was noted.463,464 The secretion of ovalbumin (a glycoprotein) from hen oviduct was not blocked by tunicamycin.465... 

Ovalbumin, a glycoprotein that is often present in standard molecular weight mixtures, gives a positive result with the Con A-HRP detection system. 

Fig. 1. SDS-PAGE pattern of human IgG heavy chain eluted from PVDF membrane. Human IgG (10 pg) was resolved by SDS-PAGE and transferred to a PVDF membrane. The proteins on the membrane were stained with amido black in water, and the heavy chain was excised and eluted with guanidinium hydrochloride/lysophosphatidylcholine. After precipitation with absolute alcohol, the glycoprotein was subjected to analytical SDS-PAGE, and the gel was stained with Coomassie brilliant blue to ascertain its purity. Lanes 1 and 4 original commercial preparation of human IgG lanes 2 and 3 IgG heavy chain eluted from the PVDF membrane lane 5 mixture of molecular mass standards, from top to bottom phosphorylase b (94 kDa), BSA (67 kDa), ovalbumin (43 kDa), carbonic anhydrase (30 kDa), soybean trypsin inhibitor (20 ldDa), ct-lactalbumin (14.4 kDa).

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. 

Rabouille, Cortassa, and Aon[81 dried protein, glycoprotein, or polysaccharide containing brine solutions that resulted in dendritic-like fractal patterns. The fractal dimension, D = 1.79, was determined for the pattern afforded by an ovomucin-ovalbumin mixture (0.1 M NaCl). Similar D values were obtained for dried solutions of fetuin, ovalbumin, albumin, and starch the authors subsequently suggest that fractal patterning is characteristic of biological polymers. 

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