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与结核分枝杆菌PE25/PPE41和PE35/PPE68相互作用的宿主蛋白的筛选 被引量:1
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作者 李田田 陈利苹 刘思国 《中国预防兽医学报》 CAS CSCD 北大核心 2017年第6期504-507,共4页
为筛选与结核分枝杆菌(MTB)的PE25/PPE41和PE35/PPE68两对异源二聚体相互作用的宿主蛋白,本研究以这两对异源二聚体为诱饵蛋白筛选与其相互作用的宿主蛋白。首先将PCR扩增的PE25/PPE41或PE35/PPE68基因串联克隆于改造的pc DNA3.1(+)中(... 为筛选与结核分枝杆菌(MTB)的PE25/PPE41和PE35/PPE68两对异源二聚体相互作用的宿主蛋白,本研究以这两对异源二聚体为诱饵蛋白筛选与其相互作用的宿主蛋白。首先将PCR扩增的PE25/PPE41或PE35/PPE68基因串联克隆于改造的pc DNA3.1(+)中(在5'端引入了以编码烟草蚀纹病毒蛋白酶切位点的序列连接的ZZ和Flag标签序列),构建诱饵重组质粒pcDNA-PE25-PPE41和pc DNA-PE35-PPE68。分别将构建的诱饵重组质粒转染293T细胞后提取细胞总蛋白,经ZZ和Flag标签蛋白两步串联亲和层析钓取宿主蛋白,SDS-PAGE电泳分离后切取差异蛋白胶条进行质谱分析,结果显示,PE25/PPE41鉴定到15个差异蛋白,PE35/PPE68鉴定到29个差异蛋白,并分别挑选了与PE25/PPE41和PE35/PPE68相互作用的3个和7个差异蛋白,为进一步验证与PE25/PPE41和PE35/PPE68直接作用的宿主蛋白及研究其生物学功能奠定了基础。 展开更多
关键词 结核分枝杆菌 pe25/ppe41 pe35/ppe68 宿主互作蛋白 串联亲和纯化
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Evolution of PE35 and PPE68 Gene Families in Mycobacterium: Roles of Horizontal Gene Transfer and Evolutionary Constraints
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作者 Ashay Bavishi Lin Lin +1 位作者 Madhusudan Choudhary Todd P. Primm 《Journal of Tuberculosis Research》 2014年第4期181-198,共18页
Mycobacterium is a genus of bacteria with over a hundred non-pathogenic and pathogenic species, best recognized for certain members known to cause diseases such as tuberculosis and leprosy. Two novel protein families ... Mycobacterium is a genus of bacteria with over a hundred non-pathogenic and pathogenic species, best recognized for certain members known to cause diseases such as tuberculosis and leprosy. Two novel protein families important in the pathogenesis of Mycobacterium species are the PE and PPE families. These two protein families affect the antigenic profiles, disturbing host immunity. To better understand the origin and evolution of these gene families and the differences in their composition between pathogenic and non-pathogenic strains, several bioinformatic analyses were conducted both among Mycobacterium and closely related species that contain PE35 and PPE68 gene homologs. The methods included protein homology searches (BLASTP), horizontal gene transfer analysis (IslandViewer), phylogenetic analysis, gene cluster analysis and structural and functional constraints. Results revealed that PE and PPE gene homologs were not only limited to Mycobacterium, but also existed in three other non-mycobacterial genera, Rhodococcus, Tsukamurella and Segniliparus, and were possibly initially acquired from non-mycobacterial microorganisms by multiple horizontal gene transfers. Results also demonstrated that PE and PPE genes were more diverse and more rapidly evolving in pathogenic Mycobacterium as compared with non-pathogenic Mycobacterium and other non-mycobacterial species. These findings possibly shed light on the diverse functions and origins of the PE/PPE proteins among these organisms. 展开更多
关键词 pe35 ppe68 HORIZONTAL GENE TRANSFER MYCOBACTERIUM
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结核分枝杆菌抗原PE35和PPE68基因多态性的初步研究 被引量:1
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作者 仇艳 蒋毅 +2 位作者 刘海灿 李马超 万康林 《疾病监测》 CAS 2015年第7期539-545,共7页
目的为研究结核分枝杆菌RD1区编码PE/PPE蛋白家族中的PE 35和PPE 68基因的多态性,并评估其基因多态性对免疫识别的影响。方法笔者从本实验室保存的来自国内不同地区的2346株结核分枝杆菌复合群临床分离株中,选取代表不同Spoligotyping... 目的为研究结核分枝杆菌RD1区编码PE/PPE蛋白家族中的PE 35和PPE 68基因的多态性,并评估其基因多态性对免疫识别的影响。方法笔者从本实验室保存的来自国内不同地区的2346株结核分枝杆菌复合群临床分离株中,选取代表不同Spoligotyping类型的161株临床分离结核分枝杆菌菌株,基因扩增后进行序列比对,比较其在T细胞抗原表位的差异。结果在161株菌株中,23株显示有PE 35基因多态性,8株有PPE 68基因多态性。在PE 35基因中,除了菌株JL06018的突变外的其余突变都导致其T细胞抗原表位的改变。PPE 68基因中,62个T细胞表位中有58个(占93.5%)发生了改变。非北京家族PE 35基因的突变率明显高于北京家族(P<0.05),而PPE 68基因突变虽是非北京家族高于北京家族,但差异无统计学意义。结论 PE 35和PPE 68的编码基因存在明显的多态性,并很有可能导致蛋白功能的明显改变。此外,这2个蛋白的人类T细胞表位是高度可变的,提示这2种蛋白可能通过参与分化选择来逃避宿主免疫。北京家族可能比非北京家族更容易被宿主T细胞识别,进而更容易传播。 展开更多
关键词 多态性 结核分枝杆菌 pe 35 ppe 68
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