SepF蛋白对猪链球菌细胞分裂的影响
发布时间：2018-11-06 07:31
【摘要】：猪链球菌(Streptococcus suis)是一种人兽共患的传染病病原,在全球有16个国家和地区都有猪链球菌病流行的报道,不仅对养猪业造成巨大的经济损失,而且严重威胁人类健康。猪链球菌根据荚膜抗原成分的不同分成33个血清型,其中猪链球菌2型(SS2)能够感染人,是流行最广、毒力最强的优势血清型。1998年和2005年中国爆发了两次由SS2感染人引起的疫情,总共造成229人发病,52人死亡的严重后果。随着抗生素的广泛使用,猪链球菌的耐药问题日益严重。病原细菌的细胞分裂蛋白质机器(protein machinary)是新的抗菌药物创制的重要靶标。与杆菌不同,链球菌具有特殊的细胞分裂机制。然而,链球菌细胞分裂机制缺乏深入研究。隔膜的形成及正确定位是细菌细胞分裂的关键环节。细菌细胞分裂骨架蛋白FtsZ与多种细胞分裂相关蛋白质相互作用形成Z环和隔膜。最近的研究发现,在枯草芽孢杆菌中存在一种膜结合蛋白SepF,可作为Z环与细胞膜之间的“桥梁”蛋白,将Z环精确的锚定到细胞中部的细胞膜上。SepF在链球菌细胞分裂过程中的功能是否与杆菌一样尚不清楚。本研究以猪链球菌为研究材料,研究SepF在链球菌生长和细胞分裂中的功能,取得的主要研究结果如下:1.SepF对SS2生长和细胞形态的影响:以SS2野生菌株SC-19为亲本菌株,构建了sepF基因缺失菌株和互补菌株。生长曲线、革兰氏染色和扫描电镜观察发现,sepF基因缺失后,SS细胞生长速度减慢,细菌链变长,透射电镜和超分辨显微镜观察发现,sepF基因缺失后SS细胞呈两头稍尖的纺锤状,细胞分裂位点的隔膜连接线与细胞纵轴不垂直、隔膜向胞内凹陷错位。可见,SepF蛋白的确影响猪链球菌的生长和细胞分裂。2.SepF蛋白与FtsZ蛋白的相互作用:细菌双杂交实验发现SepF蛋白与FtsZ蛋白能够互作。分别克隆、表达和纯化SepF和FtsZ蛋白,体外实验也证实了二者的相互作用,并且SepF能够促进FtsZ蛋白的聚合形成多聚体。此外还发现SepF蛋白能与肽聚糖合成酶MurG互作。这些结果预示与杆菌一样,SepF蛋白可能在链球菌中也协助正确选择细胞分裂位点,影响细胞分裂中隔膜形态,在维持细胞形态具有重要功能。
[Abstract]:Streptococcus suis (Streptococcus suis) is a zoonotic infectious disease, which is reported in 16 countries and regions in the world, which not only causes huge economic loss to pig industry, but also seriously threatens human health. Streptococcus suis is divided into 33 serotypes according to the different components of capsule antigen. Among them, Streptococcus suis type 2 (SS2) can infect people and is the most prevalent. The most virulent predominant serotype. In 1998 and 2005, there were two outbreaks in China caused by SS2 infection, resulting in 229 cases and 52 deaths. With the widespread use of antibiotics, the drug resistance of Streptococcus suis is becoming more and more serious. (protein machinary), the cell division protein machine of pathogenic bacteria, is an important target for the creation of new antimicrobial agents. Unlike Bacillus, Streptococcus has a special cell division mechanism. However, the mechanism of streptococcus cell division is not well understood. The formation and correct location of diaphragm is the key link of bacterial cell division. Bacterial mitotic cytoskeleton protein (FtsZ) interacts with a variety of cell division related proteins to form Z rings and membranes. Recent studies have found that there is a membrane binding protein SepF, in Bacillus subtilis that acts as a "bridge" protein between Z ring and cell membrane. The Z loop was precisely anchored to the cell membrane in the middle of the cell. It was not clear whether SepF played the same role in the cell division of streptococcus as that of the bacillus. The function of SepF in the growth and cell division of Streptococcus suis was studied in this study. The main results were as follows: the effect of 1.SepF on the growth and cell morphology of SS2: the wild SS2 strain SC-19 was used as parent strain. SepF gene deletion strain and complementary strain were constructed. Growth curve, Gram staining and scanning electron microscopy showed that after sepF gene deletion, the growth rate of SS cells slowed down and the bacterial chain became longer. After the deletion of sepF gene, the SS cells were spindle-like at both ends, the connective line of the cell division site was not perpendicular to the longitudinal axis of the cell, and the diaphragm was dislocated into the cell. The interaction between 2.SepF protein and FtsZ protein was observed. The results showed that SepF protein and FtsZ protein could interact with each other. SepF and FtsZ proteins were cloned, expressed and purified, and their interaction was confirmed by experiments in vitro, and SepF promoted the polymerization of FtsZ proteins to form polymers. It was also found that SepF protein could interact with peptidoglycan synthase MurG. These results suggest that, like bacillus, SepF protein may also assist in the correct selection of cell division sites in streptococcus, affect the membrane morphology in cell division, and play an important role in maintaining cell morphology.
【学位授予单位】：华中农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S852.611
本文编号：2313571
[Abstract]:Streptococcus suis (Streptococcus suis) is a zoonotic infectious disease, which is reported in 16 countries and regions in the world, which not only causes huge economic loss to pig industry, but also seriously threatens human health. Streptococcus suis is divided into 33 serotypes according to the different components of capsule antigen. Among them, Streptococcus suis type 2 (SS2) can infect people and is the most prevalent. The most virulent predominant serotype. In 1998 and 2005, there were two outbreaks in China caused by SS2 infection, resulting in 229 cases and 52 deaths. With the widespread use of antibiotics, the drug resistance of Streptococcus suis is becoming more and more serious. (protein machinary), the cell division protein machine of pathogenic bacteria, is an important target for the creation of new antimicrobial agents. Unlike Bacillus, Streptococcus has a special cell division mechanism. However, the mechanism of streptococcus cell division is not well understood. The formation and correct location of diaphragm is the key link of bacterial cell division. Bacterial mitotic cytoskeleton protein (FtsZ) interacts with a variety of cell division related proteins to form Z rings and membranes. Recent studies have found that there is a membrane binding protein SepF, in Bacillus subtilis that acts as a "bridge" protein between Z ring and cell membrane. The Z loop was precisely anchored to the cell membrane in the middle of the cell. It was not clear whether SepF played the same role in the cell division of streptococcus as that of the bacillus. The function of SepF in the growth and cell division of Streptococcus suis was studied in this study. The main results were as follows: the effect of 1.SepF on the growth and cell morphology of SS2: the wild SS2 strain SC-19 was used as parent strain. SepF gene deletion strain and complementary strain were constructed. Growth curve, Gram staining and scanning electron microscopy showed that after sepF gene deletion, the growth rate of SS cells slowed down and the bacterial chain became longer. After the deletion of sepF gene, the SS cells were spindle-like at both ends, the connective line of the cell division site was not perpendicular to the longitudinal axis of the cell, and the diaphragm was dislocated into the cell. The interaction between 2.SepF protein and FtsZ protein was observed. The results showed that SepF protein and FtsZ protein could interact with each other. SepF and FtsZ proteins were cloned, expressed and purified, and their interaction was confirmed by experiments in vitro, and SepF promoted the polymerization of FtsZ proteins to form polymers. It was also found that SepF protein could interact with peptidoglycan synthase MurG. These results suggest that, like bacillus, SepF protein may also assist in the correct selection of cell division sites in streptococcus, affect the membrane morphology in cell division, and play an important role in maintaining cell morphology.
【学位授予单位】：华中农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S852.611
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