DAB₃₈₉IL‐2 (Denileukin diftitox) is considered an immunotoxin, and it is the first immunotoxin approved by Food and Drug Administration. It is used for the treatment of a cutaneous form of T‐cell lymphoma. This fusion protein has two disulfide bonds in its structure that play an essential role in toxicity and functionality of the immunotoxin. Escherichia coli (E. coli ) strain BL21 (DE3) is not capable of making disulfide bonds in its reductive cytoplasm, but the E. coli strain Rosetta‐gami (DE3) is a proper strain for the correct expression of the protein due to mutations in glutaredoxin reductase and thioredoxin reductase. In this study, a pET21a vector with the His6‐tag fused at the N‐terminus of DAB₃₈₉IL‐2 was used to express the soluble immunotoxin in E. coli Rosetta‐gami (DE3). After the purification of the soluble protein by two‐step column chromatographies, the structure of DAB₃₈₉IL‐2 was analyzed using the Native‐PAGE and circular dichroism methods. In the following, the nuclease activity of soluble DAB₃₈₉IL‐2 and its cytotoxicity activity were determined. It is concluded that the soluble recombinant protein expressed in the E. coli Rosetta‐gami (DE3) has an intact structure and also functional; hence, this form of immunotoxin could be competitive with its commercial counterparts.

中文翻译：

---

可溶性和功能性融合蛋白DAB389 IL-2在大肠杆菌菌株Rosetta-gami（DE3）中的表达和纯化。

DAB ₃₈₉ IL-2（Denileukin diftitox）被认为是一种免疫毒素，它是食品和药物管理局批准的第一种免疫毒素。它用于治疗皮肤形式的T细胞淋巴瘤。该融合蛋白在其结构中具有两个二硫键，它们在免疫毒素的毒性和功能性中起着至关重要的作用。大肠杆菌（大肠杆菌）菌株BL21（DE3）是不能够在其还原性细胞质使二硫键的，但大肠杆菌菌株罗塞塔-加米（DE3）是用于所述蛋白质的表达正确适当的应变由于基因突变在戊二醛还原酶和硫氧还蛋白还原酶中。在这项研究中，pET21a载体带有His6标签，融合在DAB ₃₈₉的N端IL-2用于在大肠杆菌Rosetta-gami（DE3）中表达可溶性免疫毒素。通过两步柱色谱法纯化可溶性蛋白后，使用Native-PAGE和圆二色谱法分析DAB ₃₈₉ IL-2的结构。接下来，确定了可溶性DAB ₃₈₉ IL-2的核酸酶活性及其细胞毒性活性。结论是，在大肠杆菌Rosetta-gami（DE3）中表达的可溶性重组蛋白具有完整的结构并且也具有功能。因此，这种形式的免疫毒素可能与其商业对手竞争。