The salt-inducible kinases (SIKs) 1 to 3, belonging to the AMPK-related kinase family, serve as master regulators orchestrating a diverse set of physiological processes such as metabolism, bone formation, immune response, oncogenesis, and cardiac rhythm. Owing to its key regulatory role, the SIK kinases have emerged as compelling targets for pharmacological intervention across a diverse set of indications. Therefore, there is interest in developing SIK inhibitors with defined selectivity profiles both to further dissect the downstream biology and for treating disease. However, despite a large pharmaceutical interest in the SIKs, experimental structures of SIK kinases are scarce. This is likely due to the challenges associated with the generation of proteins suitable for structural studies. By adopting a rational approach to construct design and protein purification, we successfully crystallized and subsequently solved the structure of SIK3 in complex with HG-9-91-01, a potent SIK inhibitor. To enable further SIK3-inhibitor complex structures we identified an antibody fragment that facilitated crystallization and enabled a robust protocol suitable for structure-based drug design. The structures reveal SIK3 in an active conformation, where the ubiquitin-associated domain is shown to provide further stabilization to this active conformation. We present four pharmacologically relevant and distinct SIK3-inhibitor complexes. These detail the key interaction for each ligand and reveal how different regions of the ATP site are engaged by the different inhibitors to achieve high affinity. Notably, the structure of SIK3 in complex with a SIK3 specific inhibitor offers insights into isoform selectivity.

中文翻译：

---

盐诱导激酶 3 与抑制剂复合物的结构揭示了结合和选择性的决定因素

盐诱导激酶 (SIK) 1 至 3 属于 AMPK 相关激酶家族，作为主调节因子协调代谢、骨形成、免疫反应、肿瘤发生和心律等多种生理过程。由于其关键的调节作用，SIK 激酶已成为跨多种适应症的药物干预的引人注目的目标。因此，人们有兴趣开发具有明确选择性特征的 SIK 抑制剂，以进一步剖析下游生物学和治疗疾病。然而，尽管制药界对 SIK 感兴趣，但 SIK 激酶的实验结构却很少。这可能是由于与适合结构研究的蛋白质的生成相关的挑战。通过采用合理的构建设计和蛋白质纯化方法，我们成功结晶并解析了SIK3与有效SIK抑制剂HG-9-91-01复合物的结构。为了进一步实现 SIK3 抑制剂复合物结构，我们鉴定了一种促进结晶的抗体片段，并实现了适合基于结构的药物设计的稳健方案。这些结构揭示了 SIK3 处于活性构象，其中泛素相关结构域被证明可以为该活性构象提供进一步的稳定性。我们提出了四种药理学相关且不同的 SIK3 抑制剂复合物。这些详细说明了每个配体的关键相互作用，并揭示了不同抑制剂如何接合 ATP 位点的不同区域以实现高亲和力。值得注意的是，SIK3 与 SIK3 特异性抑制剂复合物的结构提供了对异构体选择性的见解。