Photooxidation of bovine neurophysin II in the presence of rose bengal.

Abstract

In order to elucidate the role of some amino acid residues in bovine neurophysin II (NP-II) for its binding ability for oxytocin and vasopressin, NP-II has been photooxidized in the presence of rose bengal and oxygen, and the relationships between the loss of hormonesbinding ability of modified protein and the destruction of photosusceptible amino acids were examined. Photooxidation of NP-II causes rapid oxidation of single methionine residue to corresponding sulphoxide followed by a slow destruction of single tyrosine residue. More prolonged irradiation causes also slight decomposition of cystine residues. The hormonesbinding ability of NP-II is almost completely retained even when the methionine residue was completely photooxidized, but is gradually decreased with the progress of the photodegradation of the tyrosine residue. The decrease in binding ability of the photooxidized protein proceeds almost identically for oxytocin and [8-arginine] vasopressin as the ligand. The binding ability is decreased to about 70% of the original when 80% of tyrosine were degraded and to about 30% of the original when the tyrosine was completely photooxidized and about 6% of cystine residues were degraded. The influence of the destruction of tyrosine residue for the loss of hormones-binding ability of protein seemed to be amplified by the subsequent photooxidation of cystine residues since there is no direct correlation between the photodestruction of cystine residues and the decrease in the binding ability. The pH binding profiles of photooxidized NP-II are found to be essentially identical with those of native protein, indicating the non-essential role of phenolic hydroxyl group of tyrosine residue of NP-II in hormones-binding process. O-Acetylation of the tyrosine residue of NP-II with N-acetylimidazole gives no significant effect on the binding ability for oxytocin or [8-arginine] vasopressin. These findings suggest that the single methionine residue of NP-II has no contribution to the binding process, while the single tyrosine residue, particularly its aromatic ring, of NP-II may participate with the binding process of the protein to both oxytocin and vasopressin with similar contribution.