Teilum Kaare, Hoch Jeffrey C, Goffin Vincent, Kinet Sandrina, Martial Joseph A, Kragelund Birthe B
Department of Protein Chemistry, Institute of Molecular Biology and Physiology, University of Copenhagen, Øster Farimagsgade 2A, DK-1353 Copenhagen K, Denmark.
J Mol Biol. 2005 Aug 26;351(4):810-23. doi: 10.1016/j.jmb.2005.06.042.
We report the solution structure of human prolactin determined by NMR spectroscopy. Our result is a significant improvement over a previous structure in terms of number and distribution of distance restraints, regularity of secondary structure, and potential energy. More significantly, the structure is sufficiently different that it leads to different conclusions regarding the mechanism of receptor activation and initiation of signal transduction. Here, we compare the structure of unbound prolactin to structures of both the homologue ovine placental lactogen and growth hormone. The structures of unbound and receptor bound prolactin/placental lactogen are similar and no noteworthy structural changes occur upon receptor binding. The observation of enhanced binding at the second receptor site when the first site is occupied has been widely interpreted to indicate conformational change induced by binding the first receptor. However, our results indicate that this enhanced binding at the second site could be due to receptor-receptor interactions or some other free energy sources rather than conformational change in the hormone. Titration of human prolactin with the extracellular domain of the human prolactin receptor was followed by NMR, gel filtration and electrophoresis. Both binary and ternary hormone-receptor complexes are clearly detectable by gel filtration and electrophoresis. The binary complex is not observable by NMR, possibly due to a dynamic equilibrium in intermediate exchange within the complex. The ternary complex of one hormone molecule bound to two receptor molecules is on the contrary readily detectable by NMR. This is in stark contrast to the widely held view that the ternary prolactin-receptor complex is only transiently formed. Thus, our results lead to improved understanding of the prolactin-prolactin receptor interaction.
我们报道了通过核磁共振光谱法测定的人催乳素的溶液结构。我们的结果在距离约束的数量和分布、二级结构的规则性以及势能方面比之前的结构有显著改进。更重要的是，该结构差异足够大，导致在受体激活机制和信号转导起始方面得出不同结论。在此，我们将未结合的催乳素结构与同源物绵羊胎盘催乳素和生长激素的结构进行比较。未结合的催乳素/胎盘催乳素以及与受体结合后的结构相似，受体结合后未发生显著的结构变化。当第一个受体位点被占据时，在第二个受体位点观察到结合增强，这一现象被广泛解释为是由于与第一个受体结合诱导的构象变化。然而，我们的结果表明，第二个位点结合增强可能是由于受体 - 受体相互作用或其他一些自由能来源，而非激素的构象变化。用人催乳素受体的细胞外结构域对人催乳素进行滴定，随后进行核磁共振、凝胶过滤和电泳分析。通过凝胶过滤和电泳可清晰检测到二元和三元激素 - 受体复合物。二元复合物无法通过核磁共振观察到，可能是由于复合物内中间交换存在动态平衡。相反，一个激素分子与两个受体分子结合形成的三元复合物很容易通过核磁共振检测到。这与广泛持有的观点，即催乳素 - 受体三元复合物只是短暂形成形成了鲜明对比。因此，我们的结果有助于更好地理解催乳素 - 催乳素受体相互作用。
