以生化方法分析肌動蛋白與後突觸質密區蛋白之交互作用

Biochemical analysis of interactions between actin and postsynaptic density

蔣玉亭(Yu-Ting Chiang)

指導教授 ： 張兗君

國立清華大學/生命科學院/分子醫學研究所/碩士(2006年)

https://doi.org/10.6843/NTHU.2006.00638

摘要

在哺乳類動物的中樞神經系統內，突觸為兩個神經系統傳遞訊號的構造及位置。緊貼在突觸的後突觸細胞膜上，有一群結構緊密，由幾十種不同種類蛋白質所組合而成的碟狀蛋白質聚合物，稱作後突觸質密區(postsynaptic density，簡稱PSD)。因其座落於樹突狀小刺的頂端，且所含蛋白質種類繁多，其功能極有可能包含了神經傳遞質受器聚合的管制，神經傳遞質受器功能的調控，以及細胞胞內訊號傳導的控制。過去研究指出，在後突觸質密區內含有肌動蛋白，以及不同種類的肌動蛋白結合蛋白，所以後突觸質密區極有可能扮演著連結細胞肌動蛋白骨架與神經傳導物質及訊號傳遞分子的角色。本篇論文主要是利用混濁度實驗、離心沉澱實驗、以及穿透式電子顯微鏡觀察等生化方法，來研究細胞肌動蛋白與後突觸質密區蛋白的相互作用。混濁度實驗的結果顯示，加入肌動蛋白會造成後突觸質密區吸光值的增加。離心沉澱實驗的結果顯示，加入肌動蛋白會促使後突觸質密區蛋白碎片的產生。進一步的經由穿透式電子顯微鏡觀察，發現這些碎片數量眾多且形狀不一。另外，加入氯化鉀以及三磷酸腺苷(adenosine triphosphate)亦會造成類似現象，但效果不若加入肌動蛋白來的顯著。因此，本研究發現，肌動蛋白有能力影響後突觸質密區蛋白的結構，使其破碎並且產生大量的蛋白質碎片。

關鍵字

肌動蛋白 ； 後突觸質密區蛋白

並列摘要

The postsynaptic density (PSD) is a compact disk-shaped structure located beneath the postsynaptic plasma membrane of synapses of the central nervous system, and is thought to play a role in neurotransmission and synaptic plasticity. It lies at the top of the spine head, and based on its location and protein composition, its functions are proposed to control neurotransmitter receptor clustering, regulate receptor function, and to mediate intracellular signaling. Past studies of PSD have shown that a substantial amount of actin is present within the PSD complex, as well as several actin-binding proteins, suggesting that PSD serves as a link between the actin cytoskeleton and neurotransmitter receptors and other signaling molecules. In this study, it was observed that addition of actin to PSD resulted in structural changes of PSD. This effect caused by actin was shown by an increase in PSD turbidity, as indicated by higher absorbance at 340nm, and by an increase in the amount of fragmented PSD particles indicated by sedimentation assays and transmission electron microscopy studies. Also, addition of salt and ATP to PSD produced similar effects, but to a lesser extent compared to that caused by the addition of actin. In conclusion, actin is able to promote PSD fragmentation.

並列關鍵字

Actin ； Postsynaptic density

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