Course and extent of variation of equine infectious anemia virus during parallel persistent infections.

Comparisons of peptide and oligonucleotide maps of glycoproteins and RNA from nine isolates of equine infectious anemia virus (EIAV) that were generated during parallel infections of two Shetland ponies revealed that each isolate was structurally unique. Each EIAV isolate contained a unique subset of variant peptides, oligonucleotides, or both, indicating that structural variation in EIAV is a random and noncumulative process and that a large spectrum of possible EIAV variants can be generated in infected animals.

Comparison of glycoproteins by two-dimensional mapping of glycosylated peptides.

We describe here a two-dimensional mapping procedure which is capable of resolving glycopeptides isolated by lectin affinity chromatography from radioiodinated tryptic digests of glycoproteins. Glycopeptide maps were successfully produced for the model proteins alpha 1-acid glycoprotein and fetuin, as well as for the two surface glycoproteins gp90 and gp45 from equine infectious anemia virus (EIAV). Differences were detected in the glycopeptide maps obtained for the gp90 and gp45 components from two antigenically distinct strains of EIAV, demonstrating the ability of this procedure to detect variations in glycosylation in closely related glycoproteins.

Reversible staining and peptide mapping of proteins transferred to nitrocellulose after separation by sodium dodecylsulfate-polyacrylamide gel electrophoresis.

We describe a staining technique, using Ponceau S in very mild conditions, by which proteins can be visualized on nitrocellulose replicas without being permanently fixed to the membrane itself, thus allowing subsequent procedures such as immunoblotting or preparative elution of the proteins to be performed. This staining technique can detect 250 to 500 ng protein, which is essentially the same sensitivity seen for Coomassie blue staining of proteins on nitrocellulose. The Ponceau S staining technique was used to locate proteins on nitrocellulose replicas for subsequent in situ radioiodination and trypsin digestion, followed by separation of the resultant digests in two-dimensional peptide analysis.

Rapid emergence of novel antigenic and genetic variants of equine infectious anemia virus during persistent infection.

Previous results from our laboratory have demonstrated that equine infectious anemia virus displays structural variations in its surface glycoproteins and RNA genome during passage and chronic infections in experimentally infected Shetland ponies (Montelaro et al., J. Biol.

Effects of temperature, pH and detergents on the molecular conformation of the enterotoxin of Clostridium perfringens.

The effects of temperature, pH and sodium dodecyl sulfate on the conformation of the enterotoxin from Clostridium perfringens type A were followed by circular dichroism in both the peptide and aromatic regions. At near-physiological conditions (35 degrees C, pH 6.7) the enterotoxin exhibited a conformation consisting of approximately 60% pleated sheet, 40% non-periodic, and essentially no helix.