Erwinia chrysanthemi asparaginase is an important drug used in cancer treatment, especially in acute lymphoblastic leukemia (ALL). Escherichia coli periplasmic space is an ideal compartment for recombinant expression of certain proteins. To efficient secretion, an appropriate signal sequence should be chosen for each protein individually. In this study, the proper signal sequences for secretory production of Erwinia asparaginase were predicted by in silico methods. Consequently, two signal peptides, OmpA and DsbA, were selected for secretory expression of the enzyme in E. coli. Asparaginase was translocated through the cytoplasmic membrane with either DsbA signal peptide or OmpA signal peptide; Using DsbA signal peptide, 5.95 units of the enzyme was obtained per milliliter of culture media, whereas OmpA signal sequence led to some amount of periplasmic expression. Our study showed that the co-translational signal peptide, DsbA, targeted the asparaginase to cell membrane more efficiently in comparison to the post-translational signal peptide, OmpA. The combination of in silico and experimental approaches provides a way to test a wide variety of signal sequences for secretory production of the enzyme in a time- and cost-effective manner. It is a fundamental step for further studies in the enzyme production process.

中文翻译：

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大肠杆菌选择信号肽在大肠杆菌中分泌表达欧文天冬酰胺酶的实验评估

菊花欧文黄曲霉天冬酰胺酶是一种用于癌症治疗的重要药物，尤其是在急性淋巴细胞白血病（ALL）中。大肠杆菌周质空间是重组表达某些蛋白质的理想区室。为了有效分泌，应分别为每种蛋白质选择合适的信号序列。在这项研究中，通过计算机方法预测了分泌型欧文氏天冬酰胺酶的适当信号序列。因此，选择了两种信号肽OmpA和DsbA在大肠杆菌中分泌表达该酶。。天冬酰胺酶与DsbA信号肽或OmpA信号肽一起通过细胞质膜转运；使用DsbA信号肽，每毫升培养基可得到5.95单位的酶，而OmpA信号序列可导致一定量的周质表达。我们的研究表明，与翻译后信号肽OmpA相比，共翻译信号肽DsbA更有效地将天冬酰胺酶靶向细胞膜。计算机技术和实验方法的结合提供了一种以时间和成本有效的方式测试用于分泌酶的多种信号序列的方法。这是酶生产过程中进一步研究的基本步骤。