【作者】 赵勇山；

【导师】 孙家钟； 张红星；

【作者基本信息】 吉林大学 ， 物理化学， 2009， 博士

【摘要】 本文采用分子力学、分子动力学和量子力学的方法,对四种蛋白体系进行了深入的理论模拟研究,主要结果如下:1.通过同源模建和分子动力学模拟方法,构建了人类酸性哺乳动物壳多糖水解酶(hAMCase)的三维结构,预测其活性区域,并在此基础上进行了与内源性和外源性抑制剂的分子对接研究。从理论上确认了对复合物形成起到重要作用的氨基酸残基,对进一步揭示hAMCase的生物学功能以及以hAMCase为靶点的抑制剂设计具有重要的意义。2.通过同源模建和分子动力学模拟的方法,给出了人类丝氨酸消旋酶(hSR)的三维结构模型及其与多肽类抑制剂结合的复合物模型,揭示了在多肽类抑制剂的R位点上引入体积较大的苯甲基更适合作为hSR的抑制剂,同时明确了hSR与此类抑制剂结合时起重要作用的氨基酸残基,为基于人类丝氨酸消旋酶三维结构的药物设计提供了重要的参考信息。3.通过同源模建和分子动力学的模拟方法,构建了人类脂肪酰胺水解酶的(hFAAH)三维结构。通过对活性位点的分析,预测了与抑制剂的结合部位,并进行了与药物二异丙酚及其结构类似物的分子对接研究。研究结果表明,hFAAH在与抑制剂的相互作用主要是范德华相互作用。另外,氨基酸残基Phe192, Ile238, Thr377, Leu380, Phe381, Phe388和Leu404与抑制剂对接过程中起主要作用。4.通过分子力学和量子力学方法,研究了Sirtuins家族蛋白酶的催化机理。计算结果表明,该家族蛋白酶的第一步催化反应为SN2反应,第二步反应为协同反应。此外,我们还研究了Sirtuins蛋白酶家族中的保守残基Phe33在催化中所起的作用,从计算结果来看,Phe33在催化过程中起到了非常重要的作用。更多还原

【Abstract】 In recently years, along with the rapid development of computer technology and computational chemistry, molecule mechanics, molecular dynamics and quantum mechanics theories play an important role in life sciences. The molecular simulations of proteins have already become a very important and active research field. By the theoretical simulation, the three dimension (3D) structure, the interactions between receptor and lingand, and protein biochemistry mechanism can be obtained. In our thesis, molecular mechanics, molecular dynamics and quantum mechanics were used to theoretically study the 3D models of four proteins and investigate their structure properties in detail. The main results are summarized as follows:1. Homology modeling and docking studies of human acid mammalian chitinaseChitinases are endo-β-1,4-N-acetylglucosaminidases that can fragment chitin and have been identified in several organisms varying from bacteria to humans. Human chitinase (hAMCase) is a functional chitinase and the experiment showed that hAMCase contributes to Th2-mediated inflammation through IL-13-dependent mechanism, and inhibition of AMCase decreases airway inflammation and airway hyper-responsiveness. Thus, hAMCase is a potential therapeutic target for anti-inflammatory therapy in Th2-mediated diseases such as asthma.By means of the homology modeling and molecular dynamics methods, the 3D structure of hAMCase is created and refined. With this model, a flexible docking study is performed and the results indicate that allosamidin is a more preferred inhibitor than NAG2, which is in good agree with the experimental facts by Yi-Te et al. From the docking studies, the important residues for binding were identified.Glu49 and Glu276 may be the key amino acids residues interacting with the substrates, and Asp192 and Trp10 may help allosamidin steadily interact with hAMCase. These results will offer further experimental studies of structure-function relationships.2. Theoretical study of human serine racemaseD-Serine occurs at high levels in the mammalian brain, higher than even some common amino acids, and it has been shown to be an endogenous ligand for the“glycine site”of N-methyl-D-aspartate (NMDA) receptors. These receptors play central roles in excitatory neurotransmission, neuronal plasticity, and learning and memory. Over activation of the NMDA receptor is proposed to be responsible for the cell death that occurs in strokes. Other studies suggest that D-serine and NMDA receptor dysfunction play a role in the pathophysiology of Alzheimer’s disease. The enzyme serine racemase has been shown to be responsible for the physiological conversion of L-serine to D-serine. The involvement of D-serine in this breadth of pathophysiological processes makes serine racemase an excellent drug target.The three dimensional structure of human serine racemase (hSR) was modeled and refined using homology modeling and molecular dynamics simulation. The binding pattern predicted by docking studies revealed that important residues interacted with the peptide inhibitors, which are Asn148, His76, Glu130 and Lys235. The module provided further refinement of the hSR/inhibitor binding interaction that may be used as a basis for new structure based design efforts.3. Homology modeling and inhibitors binding study of human fatty acid amide hydrodaseFatty acid amide hydrolase (FAAH) is responsible for the hydrolysis of the fatty acid amide class of lipid transmitters which include the endogenous cannabinoid N-arachidonoyl ethanolamine, sleep-inducing agent oleamide, anorexigenic agent N-oleoyl ethanolamide and anti-inflammatory agent N-palmitoyl ethanolamide. Studies of FAAH inactivation suggest that this enzyme could be an attractive target for the treatment of pain, inflammatory, or sleep disorders.With the aid of homology modeling and molecular dynamics methods, the 3D model of hFAAH is constructed. The docking study is performed on the basis of propofol and its structural analogs. The results indicate the residues Phe192, Ile238, Thr377, Leu380, Phe381, Phe388 and Leu404 in hFAAH are seven important determinant residues in binding as they have strong van der Waals interactions with the inhibitors.4. Theoretical studies on the reaction mechanism catalyzed by SirtuinsSirtuins comprise a broadly conserved family of NAD+-dependent protein deacetylases and mono ADP-ribosyltransferases whose functionis crucial to the apoptosis and cell survival, transcriptional silencing, neurodegeneration and calorie restriction. As a key player in a broad variety of biological processes, sirtuins may be attractive targets for treatment of cardiovascular disease, diabetes, cancers and the aging. A thorough understanding of the catalytic mechanism of these enzymes will facilitate the development specific, mechanism-based regulators of sirtuins activity in order to treat the diseases related with these enzymes.Using molecular mechanism and quantum mechanism methods, the reaction mechanism catalyzed by Sirtuins were theoretically studied. The calculated results indicate that the fist step take places in a concerted SN2 step. The second step of the catalytic mechanism was further explored and confirmed that subsequent proton is abstracted from 2’ hydroxyl of N-ribose to His116 coupled with proton transferred from 2’ hydroxyl to 3’ hydroxyl, which is concerted reaction. In addition, we also examine the role of Phe33 in the protein structure, and the results of the calculations confirm that the Phe33 plays an important role in the first step of catalytic mechanism, which positions above the ribose oxygen adjacent to the ribose C1’ position to prevent nicotinamide exchange reaction.更多还原