Shimoji Yoshihiro, Ogawa Yohsuke, Osaki Makoto, Kabeya Hidenori, Maruyama Soichi, Mikami Takeshi, Sekizaki Tsutomu
National Institute of Animal Health, Tsukuba, Ibaraki, Japan.
J Bacteriol. 2003 May;185(9):2739-48. doi: 10.1128/JB.185.9.2739-2748.2003.
Erysipelothrix rhusiopathiae is a gram-positive bacterium that causes erysipelas in animals and erysipeloid in humans. We found two adhesive surface proteins of E. rhusiopathiae and determined the nucleotide sequences of the genes, which were colocalized and designated rspA and rspB. The two genes were present in all of the serovars of E. rhusiopathiae strains examined. The deduced RspA and RspB proteins contain the C-terminal anchoring motif, LPXTG, which is preceded by repeats of consensus amino acid sequences. The consensus sequences are composed of 78 to 92 amino acids and repeat 16 and 3 times in RspA and RspB, respectively. Adhesive surface proteins of other gram-positive bacteria, including Listeria monocytogenes adhesin-like protein, Streptococcus pyogenes protein F2 and F2-like protein, Streptococcus dysgalactiae FnBB, and Staphylococcus aureus Cna, share the same consensus repeats. Furthermore, the N-terminal regions of RspA and RspB showed characteristics of the collagen-binding domain that was described for Cna. RspA and RspB were expressed in Escherichia coli as histidine-tagged fusion proteins and purified. The recombinant proteins showed a high degree of capacity to bind to polystyrene and inhibited the binding of E. rhusiopathiae onto the abiotic surface in a dose dependent manner. In a solid-phase binding assay, both of the recombinant proteins bound to fibronectin, type I and IV collagens, indicating broad spectrum of their binding ability. It was suggested that both RspA and RspB were exposed on the cell surface of E. rhusiopathiae, as were the bacterial cells agglutinated by the anti-RspA immunoglobulin G (IgG) and anti-RspB IgG. RspA and RspB were present both in surface-antigen extracts and the culture supernatants of E. rhusiopathiae Fujisawa-SmR (serovar 1a) and SE-9 (serovar 2). The recombinant RspA, but not RspB, elicited protection in mice against experimental challenge. These results suggest that RspA and RspB participate in initiation of biofilm formation through their binding abilities to abiotic and biotic surfaces.
猪红斑丹毒丝菌是一种革兰氏阳性菌，可引起动物丹毒和人类类丹毒。我们发现了猪红斑丹毒丝菌的两种黏附表面蛋白，并确定了这两个基因的核苷酸序列，它们位于同一位置，分别命名为rspA和rspB。这两个基因存在于所检测的猪红斑丹毒丝菌菌株的所有血清型中。推导的RspA和RspB蛋白含有C末端锚定基序LPXTG，其前面是共有氨基酸序列的重复序列。共有序列由78至92个氨基酸组成，在RspA和RspB中分别重复16次和3次。其他革兰氏阳性菌的黏附表面蛋白，包括单核细胞增生李斯特菌黏附素样蛋白、化脓性链球菌蛋白F2和F2样蛋白、停乳链球菌FnBB以及金黄色葡萄球菌Cna，都具有相同的共有重复序列。此外，RspA和RspB的N末端区域显示出Cna所描述的胶原结合结构域的特征。RspA和RspB在大肠杆菌中作为组氨酸标签融合蛋白表达并纯化。重组蛋白显示出高度的结合聚苯乙烯的能力，并以剂量依赖的方式抑制猪红斑丹毒丝菌与非生物表面的结合。在固相结合试验中，两种重组蛋白都能与纤连蛋白、I型和IV型胶原结合，表明它们具有广泛的结合能力。有人提出，RspA和RspB都暴露在猪红斑丹毒丝菌的细胞表面，抗RspA免疫球蛋白G（IgG）和抗RspB IgG凝集的细菌细胞也是如此。RspA和RspB存在于猪红斑丹毒丝菌藤泽-SmR（血清型1a）和SE-9（血清型2）的表面抗原提取物和培养上清液中。重组RspA而非RspB能使小鼠免受实验性攻击。这些结果表明，RspA和RspB通过它们与非生物和生物表面的结合能力参与生物膜形成的起始过程。
