Targeted tyrosine iodination in a multi-tyrosine vasopressin analog.
Iodination of the conserved 2-tyrosine (Tyr(2)) 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 Tyr(2) 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 Tyr(2) by capping this residue with a reversible O-acetyl group, incorporated during peptide synthesis. The O-acetyl-Tyr(2) LVP peptide, with a free iodinatable tyrosine attached to the epsilon-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 epsilon-amine of LVP. Hitherto, this (125)I 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 Tyr(2), typical of vasopressin and oxytocin agonists.
Durr
Jacques A
JA
Bay Pines VA Health Care System, Bay Pines, Florida 33744, USA. jadurr@att.net
Blankenship
Mary
M
Chauhan
Satendra S
SS
Pennington
Michael W
MW
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D054879:Halogenation; D006801:Humans; D007457:Iodine Radioisotopes / Q000737:chemistry; D008969:Molecular Sequence Data; D010121:Oxytocin / Q000737:chemistry; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry; D014443:Tyrosine / Q000737:chemistry*; D014667:Vasopressins / Q000737:chemistry*