Yamamoto K, Sun W Y, Ohta M, Hamada K, DeLuca H F, Yamada S
Institute for Medical and Dental Engineering, Tokyo Medical and Dental University, Japan.
J Med Chem. 1996 Jul 5;39(14):2727-37. doi: 10.1021/jm9600048.
Two proteins play important roles in the expression of vitamin D function: the specific nuclear receptor protein (vitamin D receptor, VDR) and the transport protein (vitamin D binding protein, DBP). This study was conducted to clarify the conformation of vitamin D responsible for binding to those proteins. For the purpose, the side chain mobility of 1,25(OH)2D3 (1) and its 20-epimer, 20-epi-1,25(OH)2D3 (2), was analyzed by a systematic conformational search. The results were depicted as a three-dimensional dot map, which indicates that the side chains of the two vitamins (1 and 2) occupy different spatial regions that are separated in two areas. We denoted these areas as A and G for 1 and EA and EG for 2. Four analogs, the diastereomers at C(20) and C(22) (3-6) of 22-methylated 1,25(OH)2D3 whose side chains were confined to occupy G, A, EA, and EG, respectively, were designed. These analogs (3-6) were synthesized efficiently by a stereoselective conjugate addition of organocuprate to steroidal E- and Z-22-en-24-ones as the key step. In binding to the VDR the affinities of the analogs (3-6) relative to 1,25-(OH)2D3 (1) were 1/60, 1/3, 20, and 1/100, respectively. These results indicate that the A region is responsible for binding of 1 to VDR and the EA region for binding of 2. Only isomer 4 showed significant affinity for DBP, indicating only the A region is responsible for binding to DBP. Thus, 5 showed clear separation of binding affinities for two proteins, VDR and DBP. Having the highest known VDR affinity, (22R)-22-methyl-20-epi-1,25(OH)2D3 (5) has potential both as a therapeutic agent and as a tool to study the molecular mechanism of vitamin D-mediated gene transcription.
两种蛋白质在维生素D功能的表达中发挥着重要作用：特异性核受体蛋白（维生素D受体，VDR）和转运蛋白（维生素D结合蛋白，DBP）。本研究旨在阐明负责与这些蛋白质结合的维生素D的构象。为此，通过系统的构象搜索分析了1,25(OH)₂D₃（1）及其20-表异构体20-表-1,25(OH)₂D₃（2）的侧链流动性。结果以三维点图表示，这表明两种维生素（1和2）的侧链占据不同的空间区域，这些区域在两个区域中是分开的。我们将这些区域分别记为1的A和G以及2的EA和EG。设计了四种类似物，即22-甲基化的1,25(OH)₂D₃在C(20)和C(22)处的非对映异构体（3 - 6），其侧链分别被限制占据G、A、EA和EG。这些类似物（3 - 6）通过将有机铜酸盐立体选择性共轭加成到甾体E-和Z-22-烯-24-酮作为关键步骤高效合成。在与VDR结合时，类似物（3 - 6）相对于1,25-(OH)₂D₃（1）的亲和力分别为1/60、1/3、20和1/100。这些结果表明A区域负责1与VDR的结合，而EA区域负责2与VDR的结合。只有异构体4对DBP表现出显著亲和力，表明只有A区域负责与DBP结合。因此，5对两种蛋白质VDR和DBP的结合亲和力表现出明显的差异。（22R）-22-甲基-20-表-1,25(OH)₂D₃（5）具有已知最高的VDR亲和力，既具有作为治疗剂的潜力，又具有作为研究维生素D介导的基因转录分子机制工具的潜力。
