Study on correlation between virulence and molecular structure of type-specific antigen of Rickettsia tsutsugamushi

恙虫病立克次体存在多种抗原变异体，我们以前的研究表明这种立克次体的抗原型与其对小鼠的毒力之间的关系。由于这种微生物的抗原性变化是由于位于立克次体表面的56-KDa蛋白的类型特异性抗原所致，因此每个菌株的毒力差异似乎取决于该蛋白的分子结构。本研究通过对纯化的56-KDa蛋白的化学分析和56-KDa蛋白基因的克隆测序，测定了来自强毒株Gilliam、Karp和Kato，以及来自低毒株Kawasaki、Kuroki和Shimokoshi的56-KDa蛋白的全分子结构，并对不同菌株之间的结构进行了比较。获得的结果如下：(1)蛋白质由521-532个氨基酸组成(相当于55308-56745个道尔顿)；(2)在N-末端识别一个由22个氨基酸组成的可能的信号肽；(3)该基因的转录受多个串联启动子的调控；(4)这些蛋白质在氨基酸序列中的同源性在60%-82%之间；(5)在分子中识别出4个可变区，跨度为16-40个氨基酸。(6)几株新分离株的56-KDa蛋白基因在限制性内切酶片段分析中表现出多态性，进一步将Gilliam、Karp和Kuroki血清型菌株分为几个亚型。

Various antigenic variants are recognized in Rickettsia tsutsugamushi, and our previous studies indicated the relationship between the antigenic type of this rickettsia and its virulence to mice. Because the antigenic variety of this microorganism is due to the type-specific antigen of 56-KDa protein located on the rickettsial surface, the difference of virulence in each strain seems to depend on the molecular structure of this protein. In this study, the whole molecular structure of 56-KDa proteins originated from high-virulent strains of Gilliam, Karp and Kato, and from low-virulent strains of Kawasaki, Kuroki and Shimokoshi, were determined by chemical analysis of purified 56-KDa proteins, and by cloning and sequencing methods of 56-KDa protein genes, and the structures were compared among the strains. The results obtained are as follows :(1) the proteins consisted of 521-532 amino acids (corresponding to 55308-56745 dalton),(2) a putative signal peptide consisting of 22 amino acids was recognized at the N-terminal end,(3) transcription of the gene is regulated by several tandem promoters,(4) the homologies of these proteins among the six strains varied from 60 to 82% in the amino acid sequences,(5) 4 variable domains with spans of 16-40 amino acids were recognized in the molecules,(6) 56-KDa protein genes from several newly isolated strains amplified by PCR method showed polymorphism in the restriction enzyme fragment analyzes, and serotype strains of Gilliam, Karp and Kuroki strains were further classified into several subtypes.