以光譜法探討DPC微胞對於人類副甲狀腺素(1-34)構型之影響

Spectroscopic Study of the Conformation of Human Parathyroid Hormone (1-34) under DPC Micellar Environment

賴韋中(Wei-Zhong Lai)

指導教授 ： 錢偉鈞

朝陽科技大學/理工學院/應用化學系/碩士(2020年)

摘要

人類副甲狀腺素(1-34)(human Parathyroid Hormone(1-34), hPTH(1-34))其主要作用為調節骨骼與腎臟中鈣磷代謝，並且增加血液中鈣的濃度，進而增加骨骼的鈣含量。週期性的給予人類副甲腺素有助於人體骨骼中鈣含量增加。人類副甲狀腺素藉由與G-蛋白偶聯受體相互作用產生效用，C端結構與受體N端細胞外結構域相互作用，然後 N端與受體跨膜結構域結合，從而激活受體。本實驗透過儀器分析探討人類副甲腺素(1-34)於Dodecylphosphocholine(DPC)微胞環境中構型之變化。紫外光/可見光(UV/VVis)光譜可初步判斷兩者產生相互作用。螢光光譜實驗發現DPC濃度增加人類副甲狀腺素(1-34)放射強度增強並產生藍位移，說明序列中Trp23由親水性環境進入到疏水性環境。脈衝式磁場梯度實驗結果顯示人類副甲狀腺素(1-34)與DPC微胞之結合常數(Ka)為4.4103 M-1，自由能(ΔG0)值為-20.9 kJ/mol，表示其相互作用為一自發反應。圓二色光譜結果發現人類副甲狀腺素(1-34)於DPC環境中α-helix結構含量隨著DPC微胞濃度增加由原本的8%上升至40%，說明DPC微胞會誘導人類副甲狀腺素(1-34)產生構型變化。核磁共振光譜顯示人類副甲腺素(1-34)於水溶液中有兩段螺旋結構，分別為Glu4至Gly14及Arg20至Asp30片段。在微胞中Gly12與Glu19產生NOE訊號誘導二片段相互靠近。人類副甲狀腺素序列中Trp23、Leu24及Leu28形成疏水面與DPC微胞相互作用，由螢光與核磁共振光譜結果推測形成之疏水面可能進入微胞疏水性中心。

關鍵字

人類副甲狀腺素(1-34) ； 十二烷基磷酸膽鹼 ； 核磁共振光譜

並列摘要

The main role of human parathyroid hormone (1-34) is to regulate calcium and phosphorus metabolism in bones and kidney, and increase the concentration of calcium in the blood, thus increasing the calcium content of bones. Periodic administration of human parathyroid hormone helps increase the calcium content in human bones. Human parathyroid hormone produces its effects by interacting with G-protein coupled receptors. The C-terminal structure interacts with the N-terminal extracellular domain of the receptor, and then the N-terminal binds to the receptor transmembrane domain, thus activating the receptor. In this experiment, through instrumental analysis, the configuration changes of human parathyroid hormone (1-34) in the DPC micellar environment were explored. UV/Vis spectrum can preliminarily judge the interaction between the PTH(1-34) and DPC. The fluorescent spectrum experiment found that the increase in DPC concentration increased the radiation intensity of human parathyroid hormone (1-34) and produced a blue shift, indicating that Trp23 in the sequence entered the hydrophobic environment from the hydrophilic environment. The results of pulsed magnetic field gradient experiments show that the binding constant (Kα) of human parathyroid hormone (1-34) and DPC micellar is 4.4103 M-1, and the free energy (ΔG0) value is -20.9 kJ/mol, indicating that the effect is a spontaneous reaction. The results of circular dichroism spectroscopy found that the content of α-helix structure of human parathyroid hormone (1-34) in the DPC environment increased from the original 8% to 40% as the concentration of DPC micellar increased, indicating that DPC micellar will induce human parathyroid the element (1-34) produces a configuration change. NMR spectroscopy showed that human parathyroid hormone (1-34) has two helical structures in aqueous solution, which are Glu4 to Gly14 and Arg20 to Asp30 fragments, respectively. In micellar, Gly12 and Glu19 produce NOE signals, which induces the two fragments to approach each other. In human parathyroid hormone sequence, Trp23, Leu24 and Leu28 form a hydrophobic surface to interact with DPC micellar, according to the results of fluorescence and nuclear magnetic resonance spectroscopy, the formed hydrophobic surface may enter the hydrophobic center of the micellar.

並列關鍵字

human parathyroid hormone(1-34) ； Dodecylphosphocholine ； NMR

參考文獻

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