Insights into Yersinia pestis biofilm development:: topology and co-interaction of Hms inner membrane proteins involved in exopolysaccharide production
Insights into Yersinia pestis biofilm development:: topology and co-interaction of Hms inner membrane proteins involved in exopolysaccharide production
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DOI：
10.1111/j.1462-2920.2007.01554.x
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发表时间：
2008-06-01
影响因子：
5.1
通讯作者：
Perry, Robert D.
中科院分区：
文献类型：
--
作者：
Bobrov, Alexander G.;Kirillina, Olga;Perry, Robert D.
首先，三个操作子hmsHFRS、hmsT和hmsP负责鼠疫耶尔森菌生物膜的形成，这对于鼠疫从跳蚤到哺乳动物的阻断性传播至关重要。本研究使用特异性抗体，在hshfrs依赖性鼠疫菌生物膜的细胞外基质中检测到聚合β -1,6- n -乙酰基-d -葡萄糖样多糖。该胞外多糖（EPS）的产生受二胍酸环化酶HmsT和EAL结构域磷酸二酯酶HmsP的正调控和负调控作用。通过拓扑预测算法和构建c端与半乳糖苷酶和碱性磷酸酶的翻译融合体，确定了Hms细胞膜蛋白可溶性片段的细胞区域化，包括HmsR的糖基转移酶结构域、HmsT的二光酸环化酶/GGDEF结构域和HmsP的磷酸二酯酶/EAL结构域。利用细菌cAMP双杂交系统检测Hms内膜蛋白的多种相互作用。生化分析证实了其中一些蛋白质之间的相互作用。我们的研究结果表明，鼠疫菌生物膜EPS的合成和调控发生在细胞质中，通过提出的Hms酶复合物。
Primarily, three operons, hmsHFRS, hmsT and hmsP, are responsible for the development of a Yersinia pestis biofilm, which is essential for blockage-dependent transmission of plague from fleas to mammals. Here, using specific antibodies, a polymeric beta-1,6-N-acetyl-D-glucosamine-like polysaccharide was detected in the extracellular matrix of hmsHFRS-dependent Y. pestis biofilm. The production of this exopolysaccharide (EPS) was controlled by diguanylate cyclase HmsT and EAL domain phosphodiesterase HmsP, acting as positive and negative regulators respectively. Cellular compartmentalization of soluble segments of Hms inner membrane proteins, including the putative glycosyltransferase domain of HmsR, the diguanylate cyclase/GGDEF domain of HmsT and the phosphodiesterase/EAL domain of HmsP, was determined by a combination of topology prediction algorithms and construction of C-terminal translational fusions with beta-galactosidase and alkaline phosphatase. Multiple interactions of Hms inner membrane proteins were detected using bacterial cAMP based two-hybrid system. Biochemical analyses confirmed some of these protein-protein interactions. Our results indicate that synthesis and regulation of the Y. pestis biofilm EPS occurs in the cytoplasm by a proposed Hms enzymatic complex.