SYNTHESIS OF REAGENTS FOR THE CONSTRUCTION OF HYPUSINE AND DEOXYHYPUSINE PEPTIDES AND THEIR APPLICATION AS PEPTIDIC ANTIGENS

Two new synthetic methods which allow access to (2S)-deoxyhypusine, na tural (2S,9R)hypusine, (2S,SS)-hypusine, and deoxyhypusine- and hypusi ne-containing peptides are described. The methods involve both the con struction of a deoxyhypusine reagent in which the alpha-nitrogen prote cting group is orthogonal to the N-7 and N-12 protecting groups and an alternate synthesis of our previous hypusine reagent, a synthesis whi ch provides for better stereochemical control at C-9. Synthetic hypusi ne and deoxyhypusine can be generated from these reagents. The hypusin e-containing hexapeptide (Cys-Thr-Gly-Hpu-His-Gly) is conjugated to ov albumin (OVA), keyhole limpet hemocyanin (KLH), and a bis-maleimide; K LH conjugates are also made with the deoxyhypusine- and lysine-contain ing hexapeptides. Monoclonal antibodies are generated to the hypusine- containing hexapeptide-OVA conjugate in mice. These are isolated and s creened against the hypusine-containing hexapeptide-KLH and hypusine-c ontaining hexapeptide-bis-maleimide conjugates, as well as against the deoxyhypusine-containing and lysine-containing hexapeptide-KLH conjug ates. These antibodies may be useful in localizing intracellular hypus ine-containing peptides as well as peptides containing hypusine analog ues.