Targeted tyrosine iodination in a multi-tyrosine vasopressin analog

Iodination of the conserved 2-tyrosine (Tyr2) residue in the pressin and tocin rings of arginine- or lysine-vasopressin (AVP or LVP), and oxytocin, respectively, impairs binding to their respective receptors. Synthetic antagonists that have their Tyr2 either replaced by another amino acid or irreversibly blocked by an O-methyl or O-ethyl ether, but have, instead, an iodinatable phenol moiety outside the pressin/tocin ring, are used for radiolabeling. We explored another approach to avoid iodinating Tyr2 by capping this residue with a reversible O-acetyl group, incorporated during peptide synthesis. The O-acetyl-Tyr2 LVP peptide, with a free iodinatable tyrosine attached to the e-amine of 8-lysine, is iodinated at a neutral pH and purified by reverse-phase high-pressure liquid chromatography (HPLC) at an acidic pH, conditions under which the O-acetyl groups are stable. Deacetylation with hydroxylamine is selective, and leaves intact the disulfide bridge. The marked shortening of the HPLC retention time after deblocking produces a chemically homogeneous label, iodinated exclusively on the free tyrosine residue attached to the e-amine of LVP. Hitherto, this 125I labeled vasopressin agonist could be obtained only in low yield, via conjugation labeling with iodinated N-t-Boc-tyrosine succinimidyl ester. This fully reversible tyrosine protection strategy does not require special equipment, and retains the conserved Tyr2, typical of vasopressin and oxytocin agonists. Copyright © 2007 European Peptide Society and John Wiley & Sons, Ltd.