嗜酸乳杆菌zrx02的plnD基因克隆及生物信息学分析

PlnD gene clone of Lactobacillus acidophilus zrx02 and bioinformatic analysis

目的:对zrx02嗜酸乳杆菌细菌素编码基因进行克隆与序列分析。方法:对分离出的菌株进行16S rRNA鉴定,并根据已报道的plnD编码产物基因设计引物,以该菌基因组DNA为模板进行PCR扩增和测序,利用Pro Param tool,TMHMM等在线软件对编码蛋白进行生物信息学分析。结果:该菌为嗜酸乳杆菌,命名为嗜酸乳杆菌zrx02,该菌含有plnD基因,通过与Gen Bank中相关的抗菌素基因进行比对,发现其基因序列与Gen Bank库中的Lactobacillus plantarum strain(WP 016526896.1)plnD gene的同源性达到100%。生物信息学分析显示该基因全长355 bp,编码92个氨基酸,相对分子量10.54 ku,理论等电点9.26,该基因编码蛋白是稳定蛋白,不具有明显的跨膜结构,二级结构主要有α-螺旋、无规则卷曲和延伸链组成,三级结构结果显示plnD蛋白模型与Streptococcus pneumoniae中的ComE同源性最高。结论:对嗜酸乳杆菌zrx02 plnD基因的克隆与分析,为进一步探究细菌素的形成机制奠定理论基础。

Objective ：Lactobacillus acidophilus produces bacteriocin, bacteriocin gene was cloned and sequence was analyzed. Methods：The isolated strain was identified by 16S rRNA.Primers were designed based on the reports plnD gene, the bacteriocin related genes of Lactobacillus acidophilus zrx02 was amplified by PCR and sequenced, then the bioinformatics were determined using online software（ Pro Param tool, TMHMM）. Results： The strain was identified as Lactobacillus acidophilus and named LactobaciUus acidophilus zrxO2.The gene essential for plnD was present in LactobaciUus acidophilus zrx02, which was naturally clustered from Lactobacillus acidophilus in GenBank, the gene sequence of the plnD gene exhibited high similarity up to 100% to that of Lactobacillus plantarum strain（ WP 016526896.1 ）plnD gene.The results of sequencing showed that the full length of the plnD gene was 355 bp, which coded for 92 amino acid residues,with molecular weight of 10.54 ku, theoretical PI of 9.26, plnD encoded protein was stable protein. In addition, there was no obvious transmembrane structure present in the encoded product.The secondary structure of protein consists of a-helix, random coil and extended strand.The protein tertiary structure exhibited high similarity to ComE from Streptococcus pneumonia. Conclusion： The cloning and analysis of plnD gene in Lactobacillus acidophilus zrx02 are helpful to further explore of bacteriocin production.