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  本文選題：銅綠假單胞菌 + 群體感應系統(tǒng)��； 參考：《西北大學》2017年碩士論文

【摘要】：銅綠假單胞菌(Pseudomonas aeruginosa,PA)是一種普遍存在的革蘭氏陰性桿菌。它對許多抗生素具有很高的內在抗藥性,是病人在醫(yī)院期間發(fā)生感染的第三大致病菌,且很難治愈。因此,尋找新的藥物作用靶點開發(fā)新型抗致病力藥物是目前研究的主要內容。群體感應(Quorum Sensing,QS)是細菌之間交流的一種方式,銅綠假單胞菌中的群體感應系統(tǒng)主要由Las,Rhl和PQS三個系統(tǒng)組成,它們控制著細菌中許多重要的功能。RsaL作為QS的抑制子,與LasR共同調控著QS中信號分子的內態(tài)平衡,同時通過調節(jié)多種基因的轉錄水平,影響著毒力、生物被膜(biofilm)等許多重要的表型。然而,其具體的功能及其調節(jié)途徑并沒有被完全闡述。本實驗利用ChIP-seq在PAO1全基因組中找到了 RsaL直接結合的兩個位點,它們分別位于PA2228/PA2229和pqsH/cdpR的基因間隔區(qū)。以前的研究表明,PqsH和CdpR都與PQS信號分子的合成有關。實驗結果顯示,在rsaL基因的敲除突變體(△rsaL)中pqsH和cdpR基因的表達水平較野生型菌株PAO1下降了很多,從而也降低了 PQS信號分子的產量。rsaL基因的突變導致了 PAO1菌體中生物被膜的降低和綠膿菌素的升高,這些表型主要是由于pqsH或cdpR轉錄水平的變化引起的。此外,我們解析了 RsaL-DNA復合物的晶體結構,發(fā)現RsaL的DNA結合結構域類似于HTH結構,這一結構在許多轉錄調節(jié)子中都是十分保守的。通過定點突變和回補實驗,我們確定了 RsaL與DNA結合時起關鍵作用的氨基酸,它們分別是Arg20,Gln27,Gln38,Gly35,Ser37 和 Ser42。本研究讓我們更清楚的理解了 RsaL復雜的調控網絡,同時也為探索QS中新的調節(jié)蛋白提供了參考和依據。
[Abstract]:Pseudomonas aeruginosa (PA) is a common gram negative bacilli. It has high intrinsic resistance to many antibiotics. It is the third general pathogen infected by the patient in hospital. It is difficult to cure. Therefore, it is currently studied to find new drug targets to develop new anti pathogenic drugs. Quorum Sensing (QS) is a way of communication between bacteria. The quorum induction system in Pseudomonas aeruginosa is composed mainly of three systems, Las, Rhl and PQS. They control many important functional.RsaL in bacteria as the inhibitor of QS, and regulate the internal equilibrium of the signal molecules in QS with LasR, and simultaneously control the internal equilibrium of the signal molecules in QS. By regulating the transcriptional level of multiple genes, it affects many important phenotypes such as virulence, biofilm (biofilm) and many other important phenotypes. However, its specific functions and its regulatory pathways have not been fully explained. This experiment uses ChIP-seq to find two direct binding sites of RsaL in the whole PAO1 genome, which are located in PA2228/PA2229 and pqsH/cdp, respectively. R gene spacers. Previous studies showed that both PqsH and CdpR were related to the synthesis of PQS signal molecules. Experimental results showed that the expression level of pqsH and cdpR genes in the rsaL gene knockout mutant (delta rsaL) decreased a lot more than that of the wild type strain, thus reducing the mutation of the.RsaL gene of the PQS signal molecule resulting in the PAO1. The decrease of biofilm and the increase of Pseudomonas aeruginosa in the bacteria are caused mainly by changes in the pqsH or cdpR transcriptional levels. In addition, we have analyzed the crystal structure of the RsaL-DNA complex and found that the DNA binding domain of RsaL is similar to the HTH structure. This structure is very conservative in the many transcriptional regulators. We have identified the key amino acids in the combination of RsaL and DNA, which are Arg20, Gln27, Gln38, Gly35, Ser37 and Ser42., which let us understand the complex regulatory network of RsaL more clearly, and provide a reference and basis for exploring the new regulator of QS in QS.
【學位授予單位】：西北大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R378

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