Suitable experimental conditions are required to characterize interferon-alpha 2b synthetic peptides

The binding and antiproliferative activities of synthetic peptides 29-35 an d 122-139 of interferon-alpha 2b, both of which contain a cysteine residue in their sequences, were studied in the presence or absence of a dissociati on medium containing mainly urea, dithiothreitol and 2-mercaptoethanol. Alt hough interferon-alpha 2b peptides either did not modify or slightly increa sed I-125-labelled interferon-alpha 2b specific binding to WISH cell-membra ne receptors in the absence of dissociation medium, significant binding inh ibition was obtained when both peptides were assayed in dissociation medium . Furthermore, also in the presence of dissociating agents, the two fragmen ts inhibited cell growth in a concentration-dependent manner, the 122-139 s equence being more effective than the 29-35 sequence. No additive effect on interferon binding and cell proliferation was observed when both peptides were added simultaneously. Results obtained after submitting peptide 122-13 9 to gel filtration or PAGE under different experimental conditions showed the presence of dimers and/or noncovalent aggregates arising from intermole cular disulfide bridges or hydrophobic interactions. Thus, our results indi cated that peptide effects on I-125-labelled interferon-alpha 2b binding an d WISH cell proliferation were dearly manifested when the amount of monomer ic species increased, showing that suitable experimental conditions should be used to study peptide behavior. The ability of both peptides to effectiv ely trigger an interferon-specific biological action, such as cell growth i nhibition, strongly suggested that 29-35 and 122-139 interferon-alpha 2b fr agments constitute the conformational epitope or mimotope that interacts wi th the cytokine-specific receptor.