南美響尾蛇毒素(Crotoxin)及其結合蛋白質之作用模式

Modes of action of crotoxin and of its binding protein crocalbin

李一良(I-Liang Lee)

指導教授 ： 曾木金

國立臺灣大學/生命科學院/生化科學研究所/博士(2008年)

https://doi.org/10.6342/NTU.2008.10655

摘要

蛇類經過長年的演化後，有許多的蛇毒蛋白質分子都具有藥效，南美響尾蛇蛇毒的主要成分crotoxin具有強烈的神經毒性，會阻斷周邊運動神經的訊息傳導分子的釋放，造成獵物四肢麻痺、窒息而死亡。Crotoxin乃是由crotoxin A（CA）及crotoxin B（CB）兩個次單元體以非共價鍵所組合而成，CA不帶有生物活性卻能使CB的毒性增加數十倍，所以一般認為CA是要彌補CB缺陷的chaperon。
對於crotoxin的研究從30年代就已經開始，但至今對分子層級的作用機制並不清楚。實驗顯示CA會出現在溶液相中，只有CB會與神經膜上受體結合，此外CA會與神經膜上受體競爭CB的結合，但生理上CB所引發的神經毒性卻大為增加，彼此間存在著矛盾。進一步的分析CA與CB兩者之間比例，發現CA對CB毒性的增強呈現兩相性之變化，CA隨著比例的增加到CA：CB為1：1時，會逐漸增強CB的生理毒性，但再增加濃度，反而會造成CB的毒性開始慢慢的降低。另外，在對crotoxin的專一性結合蛋白質crocalbin的研究中，意外的發現crocalbin也可以增加CB的毒性。這些實驗在在的顯示出crotoxin與受體的結合反應十分特殊。
在建立crotoxin與神經受體的結合模型的各種參數資料後，輪廓漸漸清晰。CA可視為CB之結合蛋白，類似受體分子。在神經膜以外的位置中，CA和非專一性受體互相競爭CB分子，但由於CA對於CB的結合能力遠大於非專一性受體，所以抑制非專一性的反應；當分子擴散到神經膜上，由於CA和專一性受體對於CB的結合能力接近，CB轉而與專一性受體結合。淨結果則是CA加速了CB在神經膜以外區域的擴散，抵達神經膜後CA的位置會由專一性受體所取代，而發揮生理之毒性。從分子模型觀之，CA其實是CB分子的高溶解性受體，協助CB到達作用位置，將其命名為semi-receptor，這種利用semi-receptor協助CB除去非專一性結合而達成專一性受體結合之機制命名為protective semi-receptor model。從crotoxin的例子來看，semi- receptor不但可以促進藥物在體內的輸送，據此增強藥物的效力，並且抑制了非專一性結合，據此抑制了藥物的副作用，所以semi-receptor可說是攜帶藥物的特殊模式。

關鍵字

南美響尾蛇毒素

並列摘要

Crotoxin from the South American rattlesnake Crotalus durissus terrificus is a potent presynaptic toxin composed of two subunits: component A and component B. Component B exhibits phospholipase A2 activity and is weakly toxic by itself, whereas no activity has been found to be associated with component A alone. In the present study, we found that component A modulates the neurotoxic effect of component B with a biphasic dose-response curve. In contrast, the phospholipase A2 activity of component B is inhibited monotonically by increasing doses of component A (IC50 ~ 115 pM). In order to resolve the seemingly contradictory observations, we propose a protective semi-receptor model, in which component A acts as a soluble, protective semi-receptor to illuminate the modulatory effect of component A on the interactions of component B with bio-membranes. This model can also account for some other data in the literature. We have also determined the equilibrium dissociation constant Kd for crotoxin subunits to be ~130 pM. Using this value and the semi-receptor model, we unambiguously measured the Kd for the binding of component B to its high-affinity receptor in the neuronal membrane to be ~1.8 nM. The present system represents the first with such a mode of action to be mathematic examination in detail. This model may have important implications in more physiological systems and may also promote to design of agents to reduce the side effects of certain pharmaceuticals.

並列關鍵字

Crotoxin

參考文獻

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