Studies have demonstrated that PE_PGRS45 is constitutively expressed under various environmental conditions (such as nutrient depletion, hypoxia, and low pH) of the in vitro growth conditions examined, indicating that PE_PGRS45 protein is critical to the basic functions of M.tuberculosis. However, there are few reports about the biochemical function and pathogenic mechanism of PE_PGRS45 protein. The fact that the gene of M. tuberculosis is not easily expressed in E. coli may be mainly due to the high content of G+C and the use of unique codons. Fusion tags are indispensable tools used to improve the soluble expression of recombinant proteins and accelerate the characterization of protein structure and function. In the present study, His6, Trx, His6-MBP were used as fusion tags, but only MBP-PE_PGRS45 formed expressed solubly. The purification using His6-MBP tag specific binding to the Ni column and is easy to separate after the tag cleavage. We used the purified PE_PGRS45 to immunize New Zealand rabbits and got the serum of anti PE_PGRS45. It was found that the titer of polyclonal antibodies against PE_PGR45 was higher than 1:256000. The result shows that the purified PE_PGRS45 can induce New Zealand rabbits to produce high titer antibodies. In conclusion, the recombinant protein PE_PGRS45 was successfully expressed in E. coli and specific antiserum was prepared, which will be followed by further evaluation of these specific antigens to develop highly sensitive and specific diagnostic tests for tuberculosis.

中文翻译：

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结核分枝杆菌PE_PGRS45基因密码子优化、可溶性表达、纯化及其多克隆抗体蛋白的制备[J].

研究表明，PE_PGRS45 在所检测的体外生长条件的各种环境条件（如营养耗尽、缺氧和低 pH 值）下组成型表达，表明 PE_PGRS45 蛋白对结核分枝杆菌的基本功能至关重要。然而，关于PE_PGRS45蛋白的生化功能和致病机制的报道较少。结核分枝杆菌基因不易在大肠杆菌中表达的事实可能主要是由于高含量的 G+C 和使用了独特的密码子。融合标签是不可或缺的工具，用于提高重组蛋白的可溶性表达，加速蛋白质结构和功能的表征。在本研究中，His6、Trx、His6-MBP 被用作融合标签，但只有 MBP-PE_PGRS45 形成可溶性表达。使用 His6-MBP 标签特异性结合到 Ni 柱的纯化，标签切割后易于分离。我们用纯化的PE_PGRS45免疫新西兰兔，得到抗PE_PGRS45的血清。结果发现抗PE_PGR45的多克隆抗体效价高于1:256000。结果表明，纯化后的PE_PGRS45可诱导新西兰兔产生高效价抗体。综上所述，制备了大肠杆菌和特异性抗血清，随后将进一步评估这些特异性抗原，以开发高灵敏度和特异性的结核病诊断试验。