Futami Junichiro, Atago Yuki, Azuma Akari, Putranto Endy Widya, Kinoshita Rie, Murata Hitoshi, Sakaguchi Masakiyo
Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan.
Departments of Cell Biology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 2-5-1 Shikatacho, Kita-ku, Okayama 700-8558, Japan.
Biochem Biophys Rep. 2016 Mar 19;6:94-100. doi: 10.1016/j.bbrep.2016.03.009. eCollection 2016 Jul.
It is now known that multicomponent protein assemblies strictly regulate many protein functions. The S100 protein family is known to play various physiological roles, which are associated with alternative complex formations. To prepare sufficient amounts of heterodimeric S100A8 and S100A9 proteins, we developed a method for bicistronic coexpression from a single-vector system using cells as a host. The complex formation between S100A8 and S100A9 appears to be dependent on the thermodynamic stability of the protein during expression. The stable S100A8/A9 heterodimer complex spontaneously formed during coexpression, and biologically active samples were purified by cation-exchange chromatography. Semi-stable homodimers of S100A8 and S100A9 were also formed when expressed individually. These results suggest that the assembly of S100 protein complexes might be regulated by expression levels of partner proteins . Because protein assembly occurs rapidly after protein synthesis, coexpression of relevant proteins is crucial for the design of multicomponent recombinant protein expression systems.
现在已知多组分蛋白质组装体严格调控许多蛋白质功能。已知S100蛋白家族发挥多种生理作用，这些作用与不同的复合物形成有关。为了制备足够量的异源二聚体S100A8和S100A9蛋白，我们开发了一种以细胞为宿主、利用单载体系统进行双顺反子共表达的方法。S100A8和S100A9之间的复合物形成似乎取决于表达过程中蛋白质的热力学稳定性。在共表达过程中自发形成了稳定的S100A8/A9异源二聚体复合物，并且通过阳离子交换色谱法纯化了具有生物活性的样品。当单独表达时，也形成了S100A8和S100A9的半稳定同源二聚体。这些结果表明，S100蛋白复合物的组装可能受伴侣蛋白表达水平的调控。由于蛋白质组装在蛋白质合成后迅速发生，相关蛋白质的共表达对于多组分重组蛋白表达系统的设计至关重要。
