Drug Discovery Research, Astellas Pharma. Inc., Tsukuba, Ibaraki, Japan.
Electronic Materials Department, Idemitsu Kosan Co., Ltd., Sodegaura, Chiba, Japan.
Commun Biol. 2021 Feb 4;4(1):159. doi: 10.1038/s42003-021-01666-5.
The structural mechanisms of single-pass transmembrane enzymes remain elusive. Kynurenine 3-monooxygenase (KMO) is a mitochondrial protein involved in the eukaryotic tryptophan catabolic pathway and is linked to various diseases. Here, we report the mammalian full-length structure of KMO in its membrane-embedded form, complexed with compound 3 (identified internally) and compound 4 (identified via DNA-encoded chemical library screening) at 3.0 Å resolution. Despite predictions suggesting that KMO has two transmembrane domains, we show that KMO is actually a single-pass transmembrane protein, with the other transmembrane domain lying laterally along the membrane, where it forms part of the ligand-binding pocket. Further exploration of compound 3 led to identification of the brain-penetrant compound, 5. We show that KMO is dimeric, and that mutations at the dimeric interface abolish its activity. These results will provide insight for the drug discovery of additional blood-brain-barrier molecules, and help illuminate the complex biology behind single-pass transmembrane enzymes.
单穿膜酶的结构机制仍难以捉摸。犬尿氨酸 3-单加氧酶（KMO）是一种参与真核色氨酸分解代谢途径的线粒体蛋白，与各种疾病有关。在这里，我们报告了哺乳动物全长结构的 KMO 以其膜嵌入形式，与化合物 3（内部鉴定）和化合物 4（通过 DNA 编码化学文库筛选鉴定）在 3.0 Å分辨率下复合。尽管预测表明 KMO 有两个跨膜结构域，但我们表明 KMO 实际上是一种单穿膜蛋白，另一个跨膜结构域沿膜侧向排列，在那里它形成配体结合口袋的一部分。对化合物 3 的进一步探索导致了穿透大脑的化合物 5 的鉴定。我们表明 KMO 是二聚体，二聚体界面的突变会使其失活。这些结果将为发现其他血脑屏障分子的药物提供启示，并有助于阐明单穿膜酶背后的复杂生物学。
