Transient expression of human intracellular proteins in human embryonic kidney (HEK) 293 cells is a reliable system for obtaining soluble proteins with biologically active conformations. Contrary to conventional concepts, we found that recombinantly expressed intracellular cancer-testis antigens (CTAs) showed frequent aggregation in HEK293 cells. Although experimental subcellular localization of recombinant CTAs displayed proper cytosolic or nuclear localization, some proteins showed aggregated particles in the cell. This aggregative property was not observed in recombinant housekeeping proteins. No significant correlation was found between the aggregative and biophysical properties, such as hydrophobicity, contents of intrinsically disordered regions and expression levels, of CTAs. These results can be explained in terms of structural instability of CTAs, which are specifically expressed in the testis and aberrantly expressed in cancer cells and function as a hub in the protein–protein network using intrinsically disordered regions. Hence, we speculate that recombinantly expressed CTAs failed to form this protein complex. Thus, unfolded CTAs formed aggregated particles in the cell.

中文翻译：

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哺乳动物细胞中重组表达的癌睾丸抗原的异常聚集特性

在人胚胎肾 (HEK) 293 细胞中瞬时表达人细胞内蛋白是获得具有生物活性构象的可溶性蛋白的可靠系统。与传统概念相反，我们发现重组表达的细胞内癌睾丸抗原 (CTA) 在 HEK293 细胞中显示出频繁的聚集。尽管重组 CTA 的实验性亚细胞定位显示出适当的胞质或核定位，但一些蛋白质在细胞中显示出聚集的颗粒。在重组管家蛋白中没有观察到这种聚集特性。CTA的疏水性、内在无序区域的含量和表达水平等聚集特性和生物物理特性之间没有显着相关性。这些结果可以用 CTA 的结构不稳定性来解释，CTA 在睾丸中特异性表达并在癌细胞中异常表达，并在使用本质上无序区域的蛋白质-蛋白质网络中充当中枢。因此，我们推测重组表达的 CTA 未能形成这种蛋白质复合物。因此，未折叠的 CTA 在细胞中形成聚集颗粒。