設計與合成十一異戊二烯焦磷酸合成酶的抑制劑及其生物活性探討

Abstract

十一異戊二烯焦磷酸酶（undecaprenyl pyrophosphate synthase, UPPs）所縮合成的55個碳產物十一異戊二烯焦磷酸（undecaprenyl pyrophosphate, UPP）在細菌細胞壁生合成過程中扮演者重要的角色。如果能有效地抑制UPPs的生物活性，進而能抑制細菌細胞壁的生合成，達到抑制細菌生長的功效。本論文的研究方向主要分為兩個部份，目的在發展可能具有抑制UPPs生物活性的小分子。 第一部分是依據UPPs和其天然受質法呢基焦磷酸（farnesyl pyrophosphate, FPP）的X-ray共結晶圖，利用苯環當作中間骨架和Cu(I)-catalyzed [3+2] cycloaddition 發展設計了L型及Y型兩種不同形狀的分子，進行生物活性的測試及探討。此外亦利用化學計算軟體Discovery Studio 1.7做L型及Y型分子和UPPs的嵌合（Docking）模擬評比篩選。計算結果顯示L型分子和目標酵素具有較好的結合能力，而所進行的酵素抑制實驗結果顯示L型分子也同樣使酵素具有較低的殘餘活性。 第二部份則是利用靶標導向合成（target-guided synthesis，TGS）的概念，依據UPPs活性中心位置的特性，設計並成功地合成出八個具有疊氮官能基團的疏水性分子片段及五個具有炔基的親水性的分子片段，以目標酵素UPPs當做模版一次進行四十組篩選，並使用質譜儀監測是否有反應的發生，篩選可能具有生物活性的前導藥物。此外也利用化學合成方法成功地合成出具有質譜訊號的化合物，進行生物活性的測試。

Undecaprenyl pyrophosphate synthase (UPPs) catalyzes eight consecutive condensation reactions of farnesyl pyrophosphate (FPP) with isopyrophosphate (IPP) to form a 55-carbon long-chain product, which is a lipid carrier for peptidoglycan synthesis of bacterial cell wall. In this study, we used two strategies to develop potential molecules which can inhibit UPPs. The first strategy is to design Y-shaped and L-shaped molecules based on the X-ray crystal structure analysis of UPPs. Both of biological assay and molecular modeling results show that L-shaped molecules have better binding affinity towards UPPs resulting in inhibitory effect. Among all the compounds investigated, L-shaped compound 22 exhibits the best inhibitory properties decrease 28 % activity of UPPs at 100 μM. The second strategy is to develop potential inhibitors by “target- guided synthesis”. It is hoped that the energy itself serves as a template to promote “1,3-dipolar“ cyclization to form the product with proper geometric and electronic fitness for the active site. To provide the necessary functionalities for the reaction and complement electronic properties of UPPs active site, two types of molecular fragments were designed and synthesized. Different combination of there two types of molecular fragments were mixed in the presence of UPPs and evaluated the possibility of forming the cycliztion compounds with the aid of eletrospray mass spectrometry.