- 學位論文
以分子動力模擬探討副甲狀腺素與副甲狀腺素受體之結構
Full atomic simulation of the parathyroid hormone/ parathyroid hormone-related protein type 1 receptor ligand binding
摘要
副甲狀腺素受體(parathyroid hormone/ parathyroid hormone-related protein type 1 receptor, PTHR1)為B族的G蛋白偶聯受體(class B family of G protein couple receptors)，其蛋白質結構由七個跨膜α螺旋所組成，在生物體內扮演調節組職生長、骨質形成、血鈣離子濃度平衡的重要腳色[1]。副甲狀腺素受體能與副甲狀腺素(parathyroid hormone, PTH)、副甲狀腺素相關蛋白(parathyroid hormone-related protein)，兩種配體結合(ligand binding)，進而改變其立體構型而傳遞信號，引起多種生物化學反應。然而目前研究對於副甲狀腺素受體的結構所知相當有限，僅有膜外(extracellular domain)的部分結構經由實驗揭露，而被認為是造成信號傳遞變化的膜內結構(transmembrane domain)則還未有可參考的實驗數據。副甲狀腺素受體與骨質疏鬆、軟骨病變、副甲狀腺功能低下等等多種疾病息息相關，若能釐清副甲狀腺素受體與配體結合的構型變化，將有助於了解這些疾病的成因，並能幫助以副甲狀腺素受體為標的之藥物結構設計(structure-based drug design)。 B族的G蛋白偶聯受體僅有升糖素受體(glucagon receptor, GCGR)、促腎上腺皮質激素釋放因子激素受體(corticotropin releasing factor receptor 1)兩者經由實驗解出膜內結構的主要部分。本研究將利用B族的G蛋白偶聯受體其膜內結構之間的相似性，參考現有的升糖素受體以及促腎上腺皮質激素釋放因子激素受體的結構，使用比較蛋白結構模型軟體(comparative protein structure modelling software) Modeller，建立副甲狀腺素受體之跨膜結構，並對其蛋白質的二級結構、極性、雙硫鍵、氫鍵，以及B族G蛋白偶聯受體的結構特色做一探討，並與文獻比較。並搭配現有實驗的副甲狀腺素膜外結構(extracellular domain)以分子動力模擬以及Modeller，建立完整的副甲狀腺素受體結構。 本研究以前述所建立的完整副甲狀腺素受體與其兩個配體，副甲狀腺素與副甲狀腺素相關蛋白在水溶液環境下分別進行分子動力模擬，以預測其構型變化以及副甲狀腺素受體與其兩個配體之間的結合點位(binding site)。因副甲狀腺素受體位於細胞膜上，為了更接近真實情況地模擬副甲狀腺素受體與副甲狀腺素、副甲狀腺素相關蛋白的交互作用，本研究提出了如何建置細胞膜環境於副甲狀腺素受體的方法，為細胞膜內的副甲狀腺素受體之跨膜結構提供了更好的分子模型，並提供建模流程方法於附錄頁。並以分子動力模擬，觀察分析副甲狀腺素受體在此環境下的變化，可以觀察到氫鍵明顯增加，整體結構比起在水溶液的環境下更趨穩定。本研究逐漸建置細胞膜環境於副甲狀腺素受體上，並利用分子模擬更進一步探討副甲狀腺素受體與副甲狀腺素以及副甲狀腺素相關蛋白的交互作用，包含結合點位、構型變化，以期能了解此一生物化學反應的機制。 本研究首先以現有的B族G蛋白偶聯受體的結構(升糖素受體、促腎上腺皮質激素釋放因子激素受體)為參考，利用比較蛋白結構軟體，Modeller以及分子模擬，預測副甲狀腺素受體之結構，並與文獻在二級結構上、雙硫鍵、以及B族G蛋白偶聯受體獨有的V-shaep現象有良好的一致性。更進一步利用分子模擬預測副甲狀腺素受體與副甲狀腺素以及副甲狀腺素相關蛋白的交互作用，探討其結合點位與構型變化。本研究以模擬觀點提供B族G蛋白偶聯受體的運作機制，更能將其結果應用於與副甲狀腺素受體相關的疾病，如原發性副甲狀腺機能亢進症(Primary hyperparathyroidism)、牙齒萌發症(Familial primary failure of tooth eruption)、E型短趾症(Brachydactyly type E)、骨質疏鬆症等多種疾病。在最後，提出與本研究相關的議題，以作為之後研究延伸的方向。
並列摘要
Parathyroid hormone/ parathyroid hormone-related protein type 1 receptor, known as PTHR1, is a family of B class G protein couple receptors, its structure contains 7 α helix. PTHR1 regulates skeletal development, bone turnover and calcium ion concentration. Binding of PTHR1 and the ligand of Parathyroid hormone, PTH and Parathyroid hormone-related protein changes the conformations of PTHR1 and plays important role in transferring the signal for crucial biochemical reactions. However, the transmembrane domain structure of PTHR1.is still unclear. Only the structure of extracellular domain of PTHR1 has been revealed by the experiments. The conformation of the transmembrane domain of PTHR1 is known to be playing important role in the signaling mechanism. On the other hand, PTHR1 is associated with many diseases, such as Osteoporosis, Hypoparathyroidism, Brachydactyl type E and Eiken syndrome. Therefore understanding and developing the full atomistic and conformation of the full length PTHR1, can provide the new insights into these diseases and help the design of drugs. The transmembrane domains of Glucagon receptor and corticotropin-releasing factor receptor 1 have been revealed by the experiments. On account of the sequence similarities, we use the comparative protein structure modelling software, Modeller to predict the transmembrane structure of PTHR1. In this thesis, we combined the molecular dynamics and Modeller to solve the structure of full length PTHR1. The hydrogen bonds, hydrophobic interactions and the disulfide bonds are analyzed to provide fundamental insights into the structure and binding sites of the PTHR1 This study also provides a framework to study the structure of PTHR1 in membrane. In the future, we can use this model to investigate the ligand binding mechanisms of PTHR1, enabling the design of new treatments with the disease for PTHR1 related diseases.
參考文獻
延伸閱讀
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