Inactivation of interferons: halomethyl ketone derivatives of phenylalanine as affinity labels

Antiviral activity of rabbit and mouse fibroblast interferons was irreversibly destroyed by treatment with halomethyl ketone derivatives of phenylalanine but not by treatment with a halomethyl ketone derivative of lysine. The inactivation reaction was pH dependent, suggesting the involvement of an amino acid residue ionizing in the region of pH 7. Tryptophan and phenylalanine, known ligands of interferons, protected rabbit interferon substantially against inactivation by the chloromethyl ketone derivative of N-tosylphenylalanine. Mixed bovine brain gangliosides protected rabbit and mouse interferons against inactivation by this reagent. Although halomethyl ketone derivatives of phenylalanine were originally designed and used for affinity labeling of the active site of chymotrypsin and similar enzymes, no evidence was found for a chymotrypsin-like activity of interferons. It is proposed that halomethyl ketone derivatives of phenylalanine inactivate interferon by an affinity labeling mechanism, first binding to a hydrophobic binding site and then reacting irreversibly with a nearby nucleophilic amino acid residue, which appears to be a histidine. This conclusion implies that a hydrophobic site on interferons is necessary for their antiviral activity.