Structure-activity relationships of neuromedin U. V. study on the stability of porcine neuromedin U-8 at the C-terminal asparagine amide under mild alkaline and acidic conditions.
Porcine neuromedin U-8 (X-Asn-NH(2), X=H-Tyr-Phe-Leu-Phe-Arg-Pro-Arg) is occasionally unstable in the biological fluids used for bioassay as well as in the acidic solutions used for purification of synthetic peptides. In this study, HPLC examination of an incubate solution of X-Asn-NH(2) revealed that the main decomposition products in Tyrode's solution (pH 7.4) were either alpha- or beta-monocarboxylic acid analogs (X-Asn-OH or X-Asp-NH(2)), and that no dicarboxylic acid analog (X-Asp-OH) was produced. Further investigation, employing a model peptide (Y-Asn-NH(2), Y=Benzoyl-Pro-Arg) incubated in a 0.1 M sodium bicarbonate solution at 60 degrees C, revealed that the decomposition of C-terminal Asn-NH(2) occurred through the formation of an aminosuccinimide intermediate (Y-Asu), at a rate faster than that of Y-Asn-Ser peptide but slower than that of Y-Asn-Gly peptide. Mild acid hydrolysis of X-Asn-NH(2) examined in a 1 M HCl solution at 60 degrees C yielded X-Asn-OH and X-Asp-NH(2), which further decomposed to yield X-Asp-OH. The C-terminal degradation of X-Asn-NH(2) resulted in reduced biological and immunochemical binding activities.
Kawai
Takako
T
Faculty of Pharmaceutical Sciences, Hokuriku University, Kanagawa-machi, Kanazawa 920-1181, Japan.
Shibata
Akira
A
Kurosawa
Katsuro
K
Sato
Yuki
Y
Kato
Sachiko
S
Ohki
Kazuhiro
K
Hashimoto
Tadashi
T
Sakura
Naoki
N
D000143:Acids; D000468:Alkalies; D000595:Amino Acid Sequence; D000818:Animals; D001216:Asparagine / Q000737:chemistry*; D002645:Chickens; D002851:Chromatography, High Pressure Liquid; D006207:Half-Life; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009119:Muscle Contraction / Q000187:drug effects; D009130:Muscle, Smooth / Q000187:drug effects; D009479:Neuropeptides / Q000737:chemistry*; D010455:Peptides / Q000138:chemical synthesis; D016339:Spectrometry, Mass, Fast Atom Bombardment; D013056:Spectrophotometry, Ultraviolet; D013329:Structure-Activity Relationship; D013388:Succinimides / Q000737:chemistry; D013552:Swine
D000143:Acids; D000468:Alkalies; D009479:Neuropeptides; D010455:Peptides; D013388:Succinimides; D001216:Asparagine; C046270:neuromedin U 8