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首页> 外文期刊>Journal of Agricultural and Food Chemistry >Cloning, Characterization, and Functional Analysis of the EPG1-2 Gene: A New Allele Coding for an Endopolygalacturonase in Kluyveromyces marxianus
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Cloning, Characterization, and Functional Analysis of the EPG1-2 Gene: A New Allele Coding for an Endopolygalacturonase in Kluyveromyces marxianus

机译:EPG1-2基因的克隆,表征和功能分析:马克斯克鲁维酵母内多聚半乳糖醛酸酶的新等位基因编码。

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摘要

A new allele, the EPG1-2 gene, which codes for an endopolygalacturonase in Kluyveromyces marxianus CECT1043, has been cloned. The gene has 1086 bp and the protein 362 amino acids, one more than the previously described Epglp. Epg1-2p shows a high degree of similarity with the polygalacturonases of fungi and yeasts. The sequences common to all of the polygalacturonases of prokaryotes, fungi, and higher plants are also conserved in Epg1-2p. The EPG1-2 gene has been expressed in Pichia pastoris, and, when fused with the signal peptide of the a-factor of Sacchar-omyces cerevisiae, the protein is properly secreted into the media. The recombinant enzyme does not appear to be fully glycosylated by P. pastoris or not glycosylated in the same manner as in K. marxianus, but it maintains the same optimum temperature (55 °C) and pH (4.5) and the same stability at different temperatures and pH values as the native enzyme, also showing the same hydrolytic behavior. The recombinant strain produces 200-fold more enzyme than the wild-type strain of K. marxianus, making it a yeast of potential industrial interest for the production of endopolygalacturonase for the food industry.
机译:已克隆了一个新的等位基因EPG1-2,该基因编码马克斯克鲁维酵母CECT1043中的内聚半乳糖醛酸酶。该基因具有1086 bp和362个氨基酸,比先前描述的Epglp多一个。 Epg1-2p与真菌和酵母的多半乳糖醛酸酶高度相似。 Epg1-2p中也保留了原核生物,真菌和高等植物的所有多半乳糖苷酶共有的序列。 EPG1-2基因已在巴斯德毕赤酵母中表达,当与酿酒酵母α-因子的信号肽融合时,该蛋白质可正确分泌到培养基中。重组酶似乎没有被巴斯德毕赤酵母完全糖基化或没有像在马克斯克鲁维酵母中一样被糖基化,但在不同温度下仍保持相同的最佳温度(55°C)和pH(4.5)和相同的稳定性温度和pH值与天然酶一样,也表现出相同的水解行为。该重组菌株产生的酶比马克斯克鲁维酵母的野生型菌株多200倍,使其成为具有潜在工业价值的酵母,可用于食品工业生产内聚半乳糖醛酸酶。

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