Murine IgE and IgG responses to melittin and its analogs.

Melittin, a bee-venom peptide of 26 amino acids, induces IgE and IgG responses in man and animals. The antibody response was shown previously to be specific primarily for the C-terminal 6 residues and its T cell epitope in H-2d restricted mice was shown to be in residue 11-19 of melittin. To study the relationship of peptide structure and immunogenicity in mice, we have prepared a series of melittin analogs varied in length and composition at the C-terminus. Immunogenicity of the analogs for IgG and IgE responses was found to correlate with two factors: a peptide length of more than 24 residues and the presence of a hydrophilic C-terminal region preferably with two to four cationic groups. These factors result in the ability of peptide to bind to cell membranes. Analogs that possess these features are good immunogens whereas those lacking any of these features are weak immunogens.

Murine T cell responses to melittin and its analogs.

The 26-residue peptide melittin from bee venom elicits high IgG1 and IgE responses in selected strains of mice. The antibody responses were shown previously to be specific mainly for the region of residue 20-26. The T cell epitope of melittin in H-2d-restricted mice is now found to be primarily in residue 11-19, corresponding to an alpha-helical amphiphilic segment of the molecule. Melittin-specific T cell lines have varying responses to different structural analogs of the melittin T cell epitope, and the results indicate that the antigenicity of T cell epitope peptides depend more on their primary structure than on their secondary structure. Melittin-specific T cell clones are found to be CD4+ and secrete IL-4, and are restricted to presentation on I-Ad or I-Ed. The I-Ad- or I-Ed-restricted clones differ in their responses to different analogs of melittin.

cDNA cloning and primary structure of a white-face hornet venom allergen, antigen 5.

A major allergen of white-face hornet (Dolichovespula maculata) venom is antigen 5 (also designated Dol m V). We have determined the primary structures of two forms of this protein by cDNA and protein sequencings. These two forms with 204 and 205 amino acid residues differ in 23% of their sequences but they are antigenically similar. Both forms have sequence similarity with a pathogenesis-related protein of tobacco leaf. In a 130-residue overlap of these proteins, 35-39 residues were identical. Hornet antigen 5 cDNAs were isolated from an expression library in lambda gt11 phage using antibody probes. Several of the cDNAs were not full length, but the fusion fragments expressed were immunoreactive. These results suggest that antigenic determinants of the sequential type are distributed throughout the entire molecule of antigen 5. After subcloning, antigen 5 was also expressed in pKK233-2 plasmid.

The photodecomposition of aminopterin.

Aminopterin is used in cell fusion experiments to select for hybrid cells by killing unfused cells which are deficient in enzymes for nucleotide salvage pathways. Aqueous aminopterin solutions exposed to fluorescent room light (wavelength greater than 300 nm) were found to lose cytotoxicity. Inactivation was accompanied by changes in the ultraviolet absorption spectra of the solutions. The ultraviolet spectrum of an irradiated aminopterin solution is used to provide a quantitative measure of its cytotoxicity. Aminopterin appears to be photolytically cleaved at the C9-N10 methylene-anilino bond in a reaction analogous to that known for the photolysis of folic acid irradiated at 365 nm (Lowry et al., 1949).