Heat-denatured Bovine Serum Albumin Research Articles

Some immunological studies with heat-denatured bovine serum albumin and its peptic, tryptic, and chymotryptic peptides.

SummaryThe effect on passive anaphylaxis and delayed hypersensitivity in guinea pigs and immune tolerance in rabbits of injections of heat-denatured bovine serum albumin (HDBSA) and its dialyzable and non-dialyzable peptides, formed by the action of crystalline trypsin, chymotrypsin, and pepsin has been presented. In some experiments, a comparison has been presented with the effect of native bovine serum albumin (BSA). A direct relationship was observed between the molecular size of the peptides and their in vivo immunological activities. There were also some differences in the immunological activity of peptides formed by the different enzymes. Chymotryptic peptides were most effective in eliciting passive anaphylaxis; whereas, there were no significant differences in the ability of nondialyzable products of various enzymes in causing Arthus and delayed hypersensitivity reactions in guinea pigs. Heat denaturation did not affect the ability of BSA to cause Arthus and delayed reactions. In fact, HDBSA and B...

Bovine serum albumin exposed to high pressure

Pressures of 1000–6000 kg/cm 2 were applied to a solution of bovine serum albumin for 15–60 min at 15–40°. The bovine serum albumin solution at pH 8.9 was compressed, for instance, at 3000 kg/cm 2 for 60 min at 35°. The product was resolved into five zones by acrylamide gel electrophoresis, and into two main and a trace boundaries by the ultracentrifuge. When results by the ultracentrifuge and those by gel electrophoresis at various gel concentrations were combined, it was found that a modified monomer, a dimer and probably higher aggregates were formed. A pressure-temperature curve was constructed; the pressure denaturation occurred in the upper region of the curve. The heat-denatured bovine serum albumin and pressure-denatured bovine serum albumin gave similar patterns in the ultracentrifuge, and gave zones with the same mobilities in gel electrophoresis. The mechanism of heat denaturation is interpreted to be identical with that of pressure denaturation, at least in the initial steps. High pressure lowered the temperature at which heat denaturation began to occur. The dimerization was irreversible. Direct measurement of the electric conductivity under high pressure revealed that another kind of aggregation, which is reversible, occurred at higher pressures than 4000 kg/cm 2 at 25–30°.

Immunochemical studies of the tryptic, chymotryptic and peptic peptides of heat denatured bovine serum albumin

An immune system, viz, heat denatured bovine serum albumin (HDBSA) and rabbit anti-HDBSA was studied to learn the nature of the antibody combining sites, Enzyme (trypsin, chymotrypsin, pepsin) digested HDBSA yielded immunologically active peptide, which were dialyzable and non-dialyzable. The peptides had molecular weights ranging from 5000 to 100,000 and showed differences in amino acid composition. The immunological activity with anti-HDBSA sera was proportional to the mol. wt. of the peptide. All active fractions, except those from peptic digests, also evoked passive cutaneous anaphylaxis (PCA) reactions in guinea pigs with antisera to native BSA. Ten to twenty times more weight of dialyzable fraction compared with non-dialyzable fraction was needed to produce equivalent inhibition of the homologous precipitin reaction and to evoke PCA reactions. Performate oxidation of HDBSA reduced immunological activity 50% and further peptic digestion abolished the ability to elicit the PCA reaction. Degradation of the trypsin resistant core of HDBSA with chymotrypsin yielded additional dialyzable peptides with molecular weights between 5000 and 10,000 having immunological activities. An immunologically active fragment of mol. wt. about 7200 was isolated from the tryptic digest. The immunological findings are consistent with the concept that HDBSA is partially extended molecule having antibody combining sites distribution among several areas on the surface rather than being restricted to one localized region.