AI-2 Mediated Quorum Sensing in Vibrio harveyi.

描述（由申请人提供）：细菌通过分泌和检测小信号分子来相互交流和协调群落的行为，这种现象被称为群体感应。提出的研究将探讨如何从细胞外信号分子的信息被转导通过膜来调节基因表达。具体来说，新发现的广泛存在的细菌AI-2自诱导剂分子与其受体LuxPQ在哈维弧菌中的相互作用将被表征。具体目标是：1。确定LuxP中影响AI-2和LuxQ相互作用的突变。将在LuxP中生成模拟ai -2结合形式的功能突变增益。显性LuxP突变也将被确定为不能与AI-2或LuxQ相互作用。2. 确定LuxQ的质周结构域中影响与LuxP相互作用的突变。基于pcr的诱变策略将用于产生增强LuxQ与LuxP相互作用的突变。在LuxQ中也将分离出抑制Aim 1中分离的LuxP的显性突变等位基因的突变。3. 测试AI-2类似物的激动剂或拮抗剂活性。将测量纯化LuxP的每个合成类似物的结合亲和力，并将其与体内AI-2活性进行比较。

DESCRIPTION (provided by applicant): Bacteria communicate with one another and coordinate the behavior of the community by means of secretion and detection of small signaling molecules, and this phenomenon is termed quorum sensing. The proposed research will investigate how the information from an extracellular signaling molecule is transduced across a membrane to regulate gene expression. Specifically, the interactions between the newly discovered and widespread bacterial AI-2 autoinducer molecule and its receptor LuxPQ in Vibrio harveyi will be characterized. The specific aims are: 1. Identify mutations in LuxP that affect interaction with AI-2 and with LuxQ. Gain of function mutations will be generated in LuxP that mimic the AI-2-bound form. Dominant LuxP mutations will also be identified that are unable to interact with AI-2 or with LuxQ. 2. Identify mutations in the periplasmic domain of LuxQ that affect interaction with LuxP. A PCR-based mutagenesis strategy will be used to generate mutations that enhance LuxQ's interaction with LuxP. Mutations will also be isolated in LuxQ that suppress dominant mutant alleles of LuxP isolated in Aim 1. 3. Test AI-2 analogs for agonist or antagonist activity. The binding affinities of each synthetic analog for purified LuxP will be measured and compared to AI-2 activity in vivo.