Pectinase enzymes from different plants have many possible biotechnological applications in various industries. Considering industrial significance of pectinolytic enzymes, the polygalacturonase (PG) gene from grapes was cloned into Escherichia coli DH5α using pTZ57R/T vector. Homologous sequences established a close link to Vitis vinifera. Conserve domain analysis predicted PLN02218 domain belongs to the cl31843 superfamily, showing its function as polygalacturonase. After confirmation by PCR and restriction analysis, the PG gene was expressed in E. coli BL21 and induced by IPTG. Expression level was assessed by 12% SDS-PAGE, which showed a 47 kDa protein. High expression level in the soluble fraction indicated that the protein is intracellular or transmembrane. Maximum expression was obtained with 1 mM IPTG and 6 hour induction time, and autoinduction with lactose increased production of the recombinant enzyme. Zymogram analysis revealed that the induced protein was an active enzyme. Expressed PG showed pectinolytic effect on the physiochemical properties of lemon juice. Natural biopolymers are greatly needed because synthetic fibers can negatively affect health. Pectin hydrolysis of banana pseudostem by the PG enzyme produced better quality fiber. Morphological studies by scanning electron microscopy revealed pectin degradation within the fiber cell architecture, showing the effectiveness of PG treatment on banana pseudostem.

中文翻译：

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葡萄酸性聚半乳糖醛酸酶的分子克隆，表征及其工业应用潜力

来自不同植物的果胶酶在各种工业中具有许多可能的生物技术应用。考虑到果胶分解酶的工业重要性，使用pTZ57R / T载体将葡萄中的聚半乳糖醛酸酶（PG）基因克隆到大肠杆菌DH5α中。同源序列建立了与葡萄的紧密联系。保守结构域分析预测PLN02218结构域属于cl31843超家族，表明其具有聚半乳糖醛酸酶的功能。经PCR和限制性酶切分析确认后，PG基因在大肠杆菌中表达BL21和IPTG诱导。通过12％SDS-PAGE评估表达水平，其显示47kDa蛋白。可溶性级分中的高表达水平表明该蛋白是细胞内或跨膜的。在1 mM IPTG和6小时的诱导时间下获得了最大的表达，而乳糖的自动诱导增加了重组酶的产量。谱图分析表明，诱导的蛋白质是一种活性酶。表达的PG表现出果胶分解作用对柠檬汁的理化性质。天然生物聚合物非常需要，因为合成纤维会对健康产生负面影响。PG酶将果胶水解香蕉假茎产生的纤维质量更好。通过扫描电子显微镜进行的形态学研究表明，果胶在纤维细胞结构中的降解，