The structures of four bombesins and their cloned precursor-encoding cDNAs from acid-solvated skin secretion of the European yellow-bellied toad, Bombina variegata

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• 作者：Bing Bai ; Hui Wang ; Yilu Xue ; Youjia Wu ; Mei Zhou ; Minjie Wei ; Tianbao Chen ; Lei Wang ; Chris Shaw
• 关键词：Amphibian ; Skin ; Peptide ; Bombesin ; Smooth muscle ; Cloning
• 刊名：Peptides
• 出版年：2012
• 出版时间：August, 2012
• 年：2012
• 卷：36
• 期：2
• 页码：221-229
• 全文大小：683 K

文摘

Four different bombesins (bombesin, His⁶-bombesin, Phe¹³-bombesin and Asp²-, Phe⁴-SAP-bombesin) have been identified by a systematic sequencing study of peptides in reverse phase HPLC fractions of the skin secretion of the European yellow-bellied toad, Bombina variegata, that had been solvated in 0.1 % (v/v) aqueous trifluoroacetic acid (TFA) and stored frozen at ?20 ¡ãC for 12 years. By using a 3¡ä- and 5¡ä-RACE PCR strategy, the corresponding biosynthetic precursor-encoding cDNAs of all four peptides were cloned from a cDNA library made from the same long-term frozen, acid-solvated skin secretion sample following thawing and lyophilization. Canonical bombesin and His⁶-bombesin are classical bombesin sub-family members, whereas Phe¹³-bombesin and Asp²-, Phe⁴-SAP-bombesin, belong to the litorin/ranatensin sub-family of bombesin-like peptides (BLPs). Assignment of these peptides to respective sub-families, was based upon both their primary structural similarities and their comparative pharmacological activities. An interesting observation in this study, was that the nucleotide sequences of the open-reading frames of cloned cDNAs encoding bombesin and its His⁶-substituted analog, were identical except for a single base that was responsible for the change observed at the position 6 residue in the mature peptide from Asn to His. In contrast, the precursor cDNA nucleotide sequences encoding the Phe¹³-bombesins, exhibited 53 base differences. The pharmacological activities of synthetic replicates of each bombesin were compared using two different mammalian smooth muscle preparations and all four peptides were found to be active. However, there were significant differences in their relative potencies.