Phosphatidylcholine-specific phospholipase C and sphingomyelinase activities in bacteria of the Bacillus cereus group
Phosphatidylcholine-specific phospholipase C and sphingomyelinase activities in bacteria of the Bacillus cereus group
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DOI：
10.1128/iai.71.11.6591-6606.2003
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发表时间：
2003-11-01
影响因子：
3.1
通讯作者：
Leppla, SH
中科院分区：
文献类型：
--
作者：
Pomerantsev, AP;Kalnin, KV;Leppla, SH
炭疽芽孢杆菌是非溶血性的，尽管它与高度溶血性蜡样芽孢杆菌密切相关。蜡样芽孢杆菌的溶血主要是由磷脂酰胆碱特异性磷脂酶 C (PC-PLC) 和鞘磷脂酶 (SPH) 的作用引起的，这两个酶分别由 plc 和 sph 基因编码。在蜡样芽孢杆菌中，这些基因被组织在一个受全局调节器 PlcR 调节的操纵子中。 B. anthracis 含有高度相似的 cereolysin 操纵子，但它在转录上是沉默的，因为 B. anthracis PlcR 在 C 末端被截短。在这里，我们报告了蜡状芽孢杆菌和炭疽芽孢杆菌的 PC-PLC 和 SPH 的克隆、表达、纯化和酶学表征。我们还研究了表达 PlcR 对 plc 和 sph 表达的影响。在蜡样芽胞杆菌中，发现 PlcR 是 plc 的正调节因子，但是 sph 的负调节因子。用来自炭疽芽孢杆菌的截短的直系同源物替换蜡样芽孢杆菌plcR基因消除了PC-PLC和SPH的活性，而将具有其自身启动子的蜡样芽孢杆菌plcR基因引入炭疽芽孢杆菌中并没有激活蜡溶菌素表达。在炭疽芽孢杆菌保护性抗原基因强启动子控制下的多拷贝质粒上含有蜡样芽孢杆菌plcR基因的炭疽芽孢杆菌菌株中或在携带含有完整蜡样芽孢杆菌plc-sph操纵子的多拷贝质粒的菌株中检测到溶血活性。当产生 PlcR 的炭疽芽孢杆菌菌株在厌氧加 CO2 或特别是在需氧加 CO2 条件下生长时，发现了轻微的溶血和 PC-PLC 激活。未修饰的炭疽芽孢杆菌亲本菌株在相同条件下没有表现出明显的溶血。
Bacillus anthracis is nonhemolytic, even though it is closely related to the highly hemolytic Bacillus cereus. Hemolysis by B. cereus results largely from the action of phosphatidyleholine-specific phospholipase C (PC-PLC) and sphingomyelinase (SPH), encoded by the plc and sph genes, respectively. In B. cereus, these genes are organized in an operon regulated by the global regulator PlcR. B. anthracis contains a highly similar cereolysin operon, but it is transcriptionally silent because the B. anthracis PlcR is truncated at the C terminus. Here we report the cloning, expression, purification, and enzymatic characterization of PC-PLC and SPH from B. cereus and B. anthracis. We also investigated the effects of expressing PlcR on the expression of plc and sph. In B. cereus, PlcR was found to be a positive regulator of plc but a negative regulator of sph. Replacement of the B. cereus plcR gene by its truncated orthologue from B. anthracis eliminated the activities of both PC-PLC and SPH, whereas introduction into B. anthracis of the B. cereus plcR gene with its own promoter did not activate cereolysin expression. Hemolytic activity was detected in B. anthracis strains containing the B. cereus plcR gene on a multicopy plasmid under control of the strong B. anthracis protective antigen gene promoter or in a strain carrying a multicopy plasmid containing the entire B. cereus plc-sph operon. Slight hemolysis and PC-PLC activation were found when PlcR-producing B. anthracis strains were grown under anaerobic-plus-CO2 or especially under aerobic-plus-CO2 conditions. Unmodified parental B. anthracis strains did not demonstrate obvious hemolysis under the same conditions.