假单胞菌M18调控因子GacA与RsmA的相关性及对抗生物质合成代谢调控机制的分析  

Analysis of mechanism and relationship of GacA and RsmA,two regulators of antibiotics production in Pseudomonas sp.M18

作　　者：葛宜和[1] 黄显清[1] 张雪洪[1] 许煜泉[1] 

机构地区：[1]上海交通大学生命科学技术学院

出　　处：《微生物学报》2006年第4期531-536,共6页Acta Microbiologica Sinica

基　　金：国家"十五"科技攻关项目(2001BA308A02-14);国家自然科学基金(30370041)~~

摘　　要：假单胞菌M18是一株可同时合成并分泌吩嗪-1-羧酸(Phenazine-1-carboxylic acid,PCA)和藤黄绿脓菌素(Pyoluteorin,Plt)两种抗生物质的生防菌株。为了进一步研究假单胞菌M18抗生物质合成代谢的调控方式与机制,在分别构建gacAr、smA等单基因突变株基础上,又构建了gacArsmA双基因突变株M18GR以及gacA′-l′acZ和rsmA′-′lacZ等翻译融合表达载体(pMEGA和pMERA)。通过在PPM和KMB两种培养基中发酵培养和两种抗生物质PCA和Plt的HPLC定量测定显示,双突变株M18GR的PCA和Plt的合成量不论在PPM还是在KMB培养基中都介于单突变株M18G和M18R之间。由实验结果分析推测,两种调控因子对抗生物质合成的调控作用不是发生在转录水平,很可能发生在转录后水平。由β-半乳糖苷酶的定量分析表明,在假单胞菌M18中,两种调控因子不存在自诱导机制;虽然GacA未调控RsmA的合成,但RsmA可能部分正向调控GacA的表达。In previous study, it has already been confirmed that the wild type strain of Pseudomonas sp. M18 isolated from the agricultural soil can produce two antifungal agents phenazine-1-carboxylic acid （PCA） and pyoluteorin （Pit）. Biosynthesis and secretion of these secondary metabolites contribute to its biological control and suppression of soilborne pathogenic fungi. As main regulators, GacA and RsmA differentially exert global regulation on production of PCA and Pit, respectively. In order to study the regulatory mechanism of secondary metabolites production in Pseudomonas sp. M18, a gacArsmA double mutant, designated as M18GR, was constructed with insertional mutation. Then, the mutant M18G, M18R, M18GR and the wild type strain M18 were inoculated into PPM or King＇s medium B （KMB）, respectively. During cultivation of strain M18 and its derivatives, their PCA and Pit were respectively detected with High Performance Liquid Chromatography （HPLC）. The results showed that PCA production in the mutant MlBGR was lower than that in the mutant MlBG and higher than that in the mutant MlBR. Pit production in the mutant MlBGR was, however, much less than that in the mutant MlBR and much more than that in the strain M18 and the mutant MlBG. With these observations, it is tempting to suggest that biosynthesis of PCA and Pit regulated by GacA or RsmA seem to occur at posttranscriptional level, not at transcriptional level. This regulation on secondary metabolites seems to be indirectly mediated by other unknown factors. Meanwhile, based on the construction of two translational fusions, gacA＇-＇lacZ and rsmA＇-＇lacZ, the assay of β-galactosidase activities in KMB medium indicated that both GacA and RsmA did not have autoinduction of their own gene expression, respectively. Although GacA did not influence expression of the rsmA gene, RsmA could exert some positive influence on the gacA gene expression in Pseudomonas sp. M18.

关 键 词：假单胞菌M18 GACA RSMA 双突变 调控机制 

分 类 号：Q936[生物学—微生物学]