Ehling-Schulz Monika, Fricker Martina, Grallert Harald, Rieck Petra, Wagner Martin, Scherer Siegfried
Lehrstuhl für Mikrobielle Okologie, Department für Biowissenschaftliche Grundlagen, WZW, Technische Universität München, Freising, Germany.
BMC Microbiol. 2006 Mar 2;6:20. doi: 10.1186/1471-2180-6-20.
Cereulide, a depsipeptide structurally related to valinomycin, is responsible for the emetic type of gastrointestinal disease caused by Bacillus cereus. Recently, it has been shown that this toxin is produced by a nonribosomal peptide synthetase (NRPS), but its exact genetic organization and biochemical synthesis is unknown.
The complete sequence of the cereulide synthetase (ces) gene cluster, which encodes the enzymatic machinery required for the biosynthesis of cereulide, was dissected. The 24 kb ces gene cluster comprises 7 CDSs and includes, besides the typical NRPS genes like a phosphopantetheinyl transferase and two CDSs encoding enzyme modules for the activation and incorporation of monomers in the growing peptide chain, a CDS encoding a putative hydrolase in the upstream region and an ABC transporter in the downstream part. The enzyme modules responsible for incorporation of the hydroxyl acids showed an unusual structure while the modules responsible for the activation of the amino acids Ala and Val showed the typical domain organization of NRPS. The ces gene locus is flanked by genetic regions with high homology to virulence plasmids of B. cereus, Bacillus thuringiensis and Bacillus anthracis. PFGE and Southern hybridization showed that the ces genes are restricted to emetic B. cereus and indeed located on a 208 kb megaplasmid, which has high similarities to pXO1-like plasmids.
The ces gene cluster that is located on a pXO1-like virulence plasmid represents, beside the insecticidal and the anthrax toxins, a third type of B. cereus group toxins encoded on megaplasmids. The ces genes are restricted to emetic toxin producers, but pXO1-like plasmids are also present in emetic-like strains. These data might indicate the presence of an ancient plasmid in B. cereus which has acquired different virulence genes over time. Due to the unusual structure of the hydroxyl acid incorporating enzyme modules of Ces, substantial biochemical efforts will be required to dissect the complete biochemical pathway of cereulide synthesis.
蜡样芽胞杆菌产生的催吐型胃肠道疾病由一种与缬氨霉素结构相关的环肽——cereulide引起。最近研究表明，这种毒素由非核糖体肽合成酶（NRPS）产生，但其确切的基因结构和生化合成过程尚不清楚。
解析了cereulide合成酶（ces）基因簇的完整序列，该基因簇编码cereulide生物合成所需的酶机制。这个24 kb的ces基因簇包含7个编码序列（CDS），除了典型的NRPS基因，如磷酸泛酰巯基乙胺基转移酶以及两个编码用于在生长的肽链中激活和掺入单体的酶模块的CDS外，上游区域还有一个编码假定水解酶的CDS，下游部分有一个ABC转运蛋白。负责掺入羟基酸的酶模块结构异常，而负责激活丙氨酸和缬氨酸的模块则显示出NRPS的典型结构域组织。ces基因座两侧是与蜡样芽胞杆菌、苏云金芽孢杆菌和炭疽芽孢杆菌的毒力质粒高度同源的基因区域。脉冲场凝胶电泳（PFGE）和Southern杂交表明，ces基因仅限于产催吐毒素的蜡样芽胞杆菌，并且确实位于一个208 kb的大质粒上，该质粒与pXO1样质粒高度相似。
位于pXO1样毒力质粒上的ces基因簇，除了杀虫毒素和炭疽毒素外，代表蜡样芽胞杆菌群在大质粒上编码的第三种毒素类型。ces基因仅限于产催吐毒素的菌株，但pXO1样质粒也存在于类似产催吐毒素的菌株中。这些数据可能表明蜡样芽胞杆菌中存在一种古老的质粒，随着时间的推移获得了不同的毒力基因。由于Ces中负责掺入羟基酸的酶模块结构异常，剖析cereulide合成的完整生化途径需要大量的生化研究工作。
