当前位置: 首页 > 文章 > 辣椒疫霉(Phytophthora capsici)多聚半乳糖醛酸酶PCIPG2N-糖基化突变体的构建与表达分析 安徽农业大学学报 2014,41 (5) 73-78
Position: Home > Articles > Construction and expression analysis of N-glycosylation mutations from PGIPG2 of Phytophthora capsici Journal of Anhui Agricultural University 2014,41 (5) 73-78

辣椒疫霉(Phytophthora capsici)多聚半乳糖醛酸酶PCIPG2N-糖基化突变体的构建与表达分析

作  者:
李艳青;夏海波;张修国
单  位:
潍坊科技学院生物工程研发中心;山东农业大学植物保护学院
关键词:
辣椒疫霉;多聚半乳糖醛酸酶PCIPG2;N-糖基化;活性
摘  要:
为研究辣椒疫霉(Phytophthora capsici)多聚半乳糖醛酸酶PCIPG2 N-糖基化对其酶活性的影响,从基因组文库中分离克隆到Pcipg2基因,利用定点突变技术突变3个潜在的糖基化位点(N34,N76,N137);构建并表达N-糖基化突变蛋白,并对突变蛋白进行温度和缓冲液体系处理检测其活性。结果发现Pcipg2基因在辣椒疫霉侵染寄主的前期表达并起重要作用。PCIPG2蛋白在30°C,pH5.0(P<0.05)的缓冲液环境下活性最高。单个的Pcipg2糖基化位点N34、N76、N137对PCIPG2的致病性起正调控作用,而3个糖基化位点相互协调的功能抑制PCIPG2的活性,在PCIPG2表达致病过程中起负调控。N-糖基化在PCIPG2酶活性上起直接作用,使得PCIPG2酶在较低水平上保持较高的稳定性。
译  名:
Construction and expression analysis of N-glycosylation mutations from PGIPG2 of Phytophthora capsici
作  者:
LI Yanqing;XIA Haibo;ZHANG Xiuguo;School of Plant Protection, Shandong Agricultural University;R&D Center for Bioengineering, Weifang University of Science &Technology;
关键词:
Phytophthora capsici;;PCIPG2;;N-glycosylation;;activity
摘  要:
The effect of N-glycosylation from polygalacturonase PCIPG2 of Phytophthora capsici was analyzed. To analyze the function of P. capsici polygalacturonases, we mutated the potential N-glycosylation sites(N34, N76 and N137) of Pcipg2(cloned the Pcipg2 gene from the genomic library) using site-directed mutagenesis. Expressed N-glycosylation proteins of mutants were treated with temperature and buffers to detect the activities. The Pcipg2 gene expressed at the early stage of Phytophthora capsici infection. The PCIPG2 protein showed the maximum activity in the buffers of pH 5 at 30°C(P<0.05). Single N-glycosylation site positively regulated the pathogenicity of PCIPG2, but the interaction of three sites inhibited the PCIPG2 activity,which negatively regulated the pathogenicity of PCIPG2. N-glycosylation can directly regulate the PCIPG2 activity to keep the PCIPG2 enzyme a high level of stability at a low level of expression.

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