单核细胞增生李斯特菌野毒株prfA基因的克隆及其分子特征分析

Cloning and molecular characteristics of prfA gene from Listeria monocytogenes wild strain

利用PCR技术对单核细胞增生李斯特菌(LM)TA野毒株prfA基因进行扩增,将其克隆后测序,并对该分离株prfA基因及其编码蛋白进行分子特征和遗传变异分析。结果显示,TA株prfA基因全长为714bp,编码237个氨基酸。通过对推导的prfA蛋白氨基酸序列结构域分析发现,该蛋白从N端到C端分别包括1个β-环结构域(18～97aa)、1个α-螺旋构成的C区(110～136aa)、1个α-螺旋构成的D区(136～155aa)、1个由螺旋-转角-螺旋构成的HTH结构域(170～196aa)和1个由α-螺旋构成的亮氨酸拉链G区结构域(211～237aa)。通过对GenBank中登录的35株分离株prfA基因遗传变异分析发现,不同地域分离株的prfA基因在核苷酸水平以点突变为主;TA株第197位氨基酸由赖氨酸(Lys,K)突变为天冬氨酸(Asn,N)。

A positive regulatory factor A（prfA） gene was amplified from Listeria monocytogenes（LM） TA isolate by PCR method and then it was cloned for sequencing.The molecular characteristics and genetic variation of prfA gene and its encoding protein were analyzed.In result,the length of prfA gene was 714 bp,encoding 237 amino acids.Protein domain analysis demonstrated that there existed a β-roll domain（18 to 97aa）,a C domain containing α-helix region（110 to 136aa）,a D domain of α-helix region（136 to 155aa）,a HTH domain of helix-turn-helix（170 to 196 aa）,and a G domain with a leucine zipper（211 to 237aa） from N-terminal to C-terminal in the deduced amino acid sequence of PrfA protein.Alignments of prfA genes of 35 LM isolates from various geographical areas indicated that site mutations were prior to other variation.The site mutation 197 of prfA gene of TA isolate resulted in the variation of amino acid from Lys to Asn.