Amphibian skin peptides and their corresponding cDNAs from single lyophilized secretion samples: Identification of novel brevinins from three species of Chinese frogs
Amphibian skin peptides and their corresponding cDNAs from single lyophilized secretion samples: Identification of novel brevinins from three species of Chinese frogs
复制标题
DOI：
10.1016/j.peptides.2005.06.024
复制
发表时间：
2006-01-01
期刊：
影响因子：
3
通讯作者：
Shaw, C
中科院分区：
文献类型：
--
作者：
Chen, TB;Li, L;Shaw, C
短杆菌肽是由24个氨基酸残基组成的肽，最初是从东方林蛙（Rana brevipoda porsa）的皮肤中分离出来的，因为它们对多种革兰氏阳性菌和革兰氏阴性菌以及致病真菌菌株具有杀菌活性。利用我们最近开发的技术，从三种未经研究的中国蛙类（花臭蛙、竹叶臭蛙和福建大头蛙）的冻干皮肤分泌物中构建了cDNA文库。在本报告中，我们描述了利用3'-RACE（快速扩增cDNA末端）技术对新型短杆菌肽进行“鸟枪法”克隆，使用了一种针对先前已鉴定的蛙皮肽cDNA的5'-非翻译区内高度保守的核酸序列区域的“通用”简并引物。从克隆的cDNA开放阅读框推导出来的新型短杆菌肽随后在相应物种皮肤分泌物的相同样本中被鉴定为成熟肽。对前原短杆菌肽核酸序列和翻译后的开放阅读框氨基酸序列进行的生物信息学分析显示出一个高度保守的信号肽区域和一个高度可变的抗菌肽编码区域。因此，这里描述的实验方法能够在不伤害供体两栖动物的情况下快速提供皮肤抗菌肽的可靠结构数据。（c）2005爱思唯尔公司。保留所有权利。
Brevinins are peptides of 24 amino acid residues, originally isolated from the skin of the Oriental frog, Rana brevipoda porsa, by nature of their microbicidal activity against a wide range of Gram-positive and Gram-negative bacteria and against strains of pathogenic fungi. cDNA libraries were constructed from lyophilized skin secretion of three, unstudied species of Chinese frog, Odorrana schmackeri, Odorrana versabilis and Pelophylax plancyi fukienensis, using our recently developed technique. In this report, we describe the "shotgun" cloning of novel brevinins by means of 3'-RACE, using a "universal" degenerate primer directed towards a highly conserved nucleic acid sequence domain within the 5'-untranslated region of previously characterized frog skin peptide cDNAs. Novel brevinins, deduced from cloned cDNA open-reading frames, were subsequently identified as mature peptides in the same samples of respective species skin secretions. Bioinformatic analysis of both prepro-brevinin nucleic acid sequences and translated open-reading frame amino acid sequences revealed a highly conserved signal peptide domain and a hypervariable anti-microbial peptide-encoding domain. The experimental approach described here can thus rapidly provide robust structural data on skin anti-microbial peptides without harming the donor amphibians. (c) 2005 Elsevier Inc. All rights reserved.