Interferon (IFN) γ can initiate immune responses by inducing the expression of major histocompatibility complex molecules, suggesting its potential for cancer immunotherapy. However, it also has an immunosuppressive function that limits its application as a therapeutic agent. IFNγ has a characteristic domain-swapped dimer structure with two of the six α-helices exchanged with each other. As we hypothesized that the contrasting functions of IFNγ could be attributed to its unique domain-swapped structure, we designed monomeric IFNγ by transforming the domain-swapped dimer structure of WT IFNγ. We conjectured the evolution of this domain-swapped dimer and hypothesized that the current IFNγ structure emerged through shortening of the loop structure at the base of the swapped domain and the accumulation of hydrophobic amino acids at the newly generated interface during domain-swapping. We then designed and generated a stable monomeric IFNγ by retracing this evolutionary process, complementing the lost loop structure with a linker, and replacing the accumulated hydrophobic amino acids with hydrophilic ones. We determined that the designed variant was a monomer based on molecular size and number of epitopes and exhibited activity in cell-based assays. Notably, the monomeric IFNγ showed a qualitatively similar balance between immunostimulatory and immunosuppressive gene expression as WT IFNγ. This study demonstrates that the structural format of IFNγ affects the strength of its activity rather than regulating the fate of downstream gene expression.

中文翻译：

---

设计单体 IFNγ：结构域交换二聚体结构在 IFNγ 免疫反应中的意义


干扰素 (IFN) γ 可以通过诱导主要组织相容性复合物分子的表达来启动免疫反应，这表明其在癌症免疫治疗中的潜力。然而，它还具有免疫抑制功能，限制了其作为治疗剂的应用。 IFNγ具有特征性的结构域交换二聚体结构，其中六个α螺旋中的两个彼此交换。由于我们假设 IFNγ 的对比功能可能归因于其独特的结构域交换结构，因此我们通过改造 WT IFNγ 的结构域交换二聚体结构设计了单体 IFNγ。我们推测了这种结构域交换二聚体的进化，并假设当前的 IFNγ 结构是通过交换结构域底部环结构的缩短以及结构域交换过程中新生成的界面处疏水性氨基酸的积累而出现的。然后，我们通过追溯这一进化过程，用连接子补充丢失的环结构，并用亲水性氨基酸取代积累的疏水性氨基酸，设计并生成了稳定的单体 IFNγ。我们根据分子大小和表位数量确定设计的变体是单体，并在基于细胞的测定中表现出活性。值得注意的是，单体 IFNγ 在免疫刺激和免疫抑制基因表达之间表现出与 WT IFNγ 类似的平衡。这项研究表明，IFNγ 的结构形式影响其活性强度，而不是调节下游基因表达的命运。