Immunogenicity Studies with Haptenated Melittin Peptides: Implication for Membrane Involvement during the Recognition Step

Abstract

Melittin peptides carrying 2,4-dinitro-6-carboxyphenyl (Dncp) haptenic groups regularly evoked anti-hapten IgG responses in mice or guinea pigs when the hapten was C-terminally attached. Single haptens on the N-terminal helix in several positions gave poor or no responses in the early stages but adequate titres after prolonged immunization. Peptides with Dncp at the C-terminus as an invariant feature and a second Dncp in various positions along the peptide chain did not fail to produce adequate responses. The hampering effect is not due to a defect at the T-cell level but involves the recognition step on the B-cell. It is implied that the haptenic interaction with the paratope of the recognizing immunoglobulin on the B-cell involves the cell membrane in an important way. It is also suggested that late antibody responses should not be overlooked during the development of proteinaceous immunogens for vaccination.

Molecular parameters in melittin immunogenicity

“…G H stands forN,N-dimethylglycin. d M±3D and the homologues M±7D, M±11D, M±15D, M±19D, M±23D as well as MD±3D with its homologues MD±7D, MD±11D, MD±15D and MD±19D have been described before [10]. located T-cell epitope must be present similar to the epitope 11±19 in H-2 d restricted mice [18].…”

“…The entire series of data is summarized in Table 2 which confirms a rather regular and unremarkably diminishing anti-Dncp IgG response of the peptides truncated stepwisw by three amino acids. Some of the antisera were also assessed against MD by immunodot assay which primarily measures antipeptidic responses [10]. Truncation has no marked effect on anti-peptide antibody generation ( Figure. 2).…”

“…GOHI-guinea pigs (out-bred, female, 250 g) from BRL, Ltd, Fu È llinsdorf, Switzerland, and Balb/c mice (female, 8±12-weeks-old) from IFFA, Credo, SaintGermain sur l'Arbresle, France, were immunized with Dncp±peptides as described before [13,10].…”

“…Solid-phase peptide synthesis according to the Fmoc/tert-butyl strategy was carried out essentially as described previously [10]. The final products were characterized by electrospray mass spectrometry and these data, together with sequences and abbreviations, are given in Table 1.…”

“…Owing to primary amphiphilicity melittin would insert inself perpendicular to the cell membrane and eventually form bundles with hydrophilic pores [6]. On the other hand the helical (secondary) amphipathy would also be expected to allow molecular positions parallel to the membrane [8,10].…”