Thyrotropin-releasing hormone (TRH) (synthetic, L-(pyro)Glu-L-His-L-Pro-NH2) and its 14C analog (L-(pyro)-Glu-L-[14C]His-L-Pro-NH2) after incubation with human plasma at 37° for 1 hr showed a 72% decrease in activity. After the recovery and purification, the peptide obtained exhibited different chromatographic and electrophoretic mobilities from TRH, but the amino acid composition was identical. The recovery of the hormone after incubation in terms of either amino acid analysis or radioactivity was 40-50%. An analogous experiment at 0° did not inactivate TRH and the recovery was 60%. The recovery of 40-50% of radioactivity from the purified Pauly-positive, inactive peptide showed that histidine is not cleaved away during incubation. Inactivated TRH did not show a free N terminus by Edmandansyl procedure suggesting that the(pyro)glutamyl ring is also intact. On treatment with [14C]diazomethane, the inactivated TRH was quantitatively converted into a [14C]methyl ester, indicating a free C terminus. Inactivated TRH was compared chromatographically and electrophoretically to synthetic (pyro)Glu-His-Pro-OH and found to have the same mobilities. These suggest that during incubation with plasma, TRH undergoes cleavage of the amide group at the prolyl end which is probably the site of inactivation.
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