Developmental expression and biochemical properties of a β-1,4-endoglucanase family in the soybean cyst nematode, Heterodera glycines
Developmental expression and biochemical properties of a β-1,4-endoglucanase family in the soybean cyst nematode, Heterodera glycines
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DOI：
10.1111/j.1364-3703.2004.00209.x
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发表时间：
2004-03-01
影响因子：
4.9
通讯作者：
Hussey, RS
中科院分区：
文献类型：
--
作者：
Gao, BL;Allen, R;Hussey, RS
大豆胞囊线虫（Heterodera glycines）产生β-1，4-内切葡聚糖酶（纤维素酶），这些酶在大豆感染期间分泌。对6个β-1，4-内切葡聚糖酶基因中的3个基因hg-eng-4、hg-eng-5和hg-eng-6的基因结构进行了分析。甘氨酸，在这里介绍。此外，我们还详细分析了β-1，4-内切葡聚糖酶基因的表达以及四种H.甘氨酸内切葡聚糖酶。其中两种内切葡聚糖酶，HG-ENG-5和HG-ENG-6，在催化结构域的氨基酸序列和基因结构上与其他四种β-1，4-内切葡聚糖酶显著不同。实时荧光定量RT-PCR结果显示，hg-eng家族成员在寄生的早期阶段有明显的发育表达差异，在寄生的后期阶段表达水平相对较低，但某些eng基因在成年雄性阶段除外。重组HG-ENG降解羧甲基纤维素，最佳酶活性范围为HG-ENG-5的pH 5.5至HG-ENG-6的pH 8。EDTA、Ca ~（2+）、Co ~（2+）、Mg ~（2+）和Fe ~（2+）对ENG蛋白的酶活性没有影响，而Zn ~（2+）、Cu ~（2+）和Mn ~（2+）对ENG蛋白的酶活性有23%~ 73%的抑制作用。在用12种不同多糖底物进行的测试中，酶活性被限制在所有测试的ENG蛋白的β-1，4键。只有HG-ENG-5和HG-ENG-6对木聚糖有较高的降解活性，对微晶纤维素有轻微的降解。总之，这些数据分别揭示了孢囊线虫寄生基因和蛋白质在表达和生物化学方面的明显差异，并揭示了寄生动物与其植物宿主之间错综复杂的相互作用。
The soybean cyst nematode, Heterodera glycines, produces beta-1,4-endoglucanases (cellulases) that are secreted during infection of soybean. The gene structures of three, hg-eng-4, hg-eng-5 and hg-eng-6, of the six beta-1,4-endoglucanase genes, all family 5 glycosyl hydrolases previously identified from H. glycines, are presented here. Furthermore, we present the detailed expression analyses of beta-1,4-endoglucanase genes as well as the biochemical properties of four H. glycines endoglucanase enzymes. Two of the endoglucanases, HG-ENG-5 and HG-ENG-6, differed significantly in their amino acid sequence of the catalytic domains and their gene structure from that of the other four beta-1,4-endoglucanases. Quantitative real-time RT-PCR revealed distinct developmental expression differences among the hg-eng family members during the early stages of parasitism and relatively low expression levels in late parasitic stages, with the exception of the adult male stage for some eng genes. Recombinant HG-ENGs degraded carboxymethylcellulose and optimum enzyme activity ranged from pH 5.5 for HG-ENG-5 to pH 8 for HG-ENG-6. EDTA, Ca2+, Co2+, Mg2+ and Fe2+ did not affect enzyme activity of any ENG protein, whereas Zn2+, Cu2+ and Mn2+ inhibited enzyme activity from 23% to 73% in some cases. In tests with 12 different polysaccharide substrates, enzyme activity was restricted to beta-1,4 linkages with all ENG proteins tested. Only HG-ENG-5 and HG-ENG-6 had relatively high activity on xylan and slightly degraded microcrystalline cellulose. Together, these data reveal distinct differences in expression and biochemistry of cyst nematode parasitism genes and proteins, respectively, and cast light on the intricate interactions between a parasitic animal and its plant host.