细胞色素p450 2s1(CYP2s1)三维结构模建及其与维甲酸分子的对接研究

利用同源模建和分子动力学模拟,模建了细胞色素P450(CYP2s1)的三维结构．在模建结构的基础上,分析了活性位点的组成和结构,并进行了与小分子(维甲酸)的分子对接研究．研究结果表明,在由维甲酸和CYP2s1形成的复合物中,非键相互作用较强,其中,GLu411和Ala414是与维甲酸相互作用能最强的两个残基,对复合物的结合起重要作用．     

Furcatin 水解酶的三维结构模建和与furcatin 的对接研究

利用同源模建和动力学模拟方法，模建了furcatin水解酶（FH）的三维结构．并在这基础上，分析了活性位点的组成和结构．研究了furcatin与FH的对接．结果表明，Ser84，Arg146，Thr189，Thr234和Gly372在复合物的形成过程中起重要的作用．其中，Ser84，Argl46和Thr189是在FH的活性口袋的二糖部分的亚单位一1中重要的氨基酸，Thr234和Gly372是亚单位-2中重要的氨基酸．     

毒蕈碱型M1受体的同源模建和分子对接

采用同源模建方法对M1受体的三维结构进行了模拟,将得到的模型分别与M受体完全激动剂乙酰胆碱和M1受体选择性激动剂占诺美林进行分子对接,形成非特异性激动和特异性激动的受体-配体复合物.用分子动力学模拟方法分别将未与小分子对接的M1受体、M1受体-乙酰且H碱复合物、M1受体-占诺美林复合物置于磷脂双膜中模拟10 ns.将模拟后的蛋白质结构与包含活性分子的测试库对接并将结果打分,以top5%富集因子(EF)作为评价依据,用占诺美林优化后的M1受体模型的EF为8.0,用乙酰胆碱优化后M1受体模型的EF为6.5,非复合物的EF为1.5.说明M1受体选择性激动剂复合物进行分子动力学模拟后得到的三维结构模型比较合理,可以作为化合物虚拟筛选的模型对新化合物进行虚拟筛选,为找到新的选择性M1受体激动剂奠定了基础.     

毒蕈碱型M1受体的同源模建和分子对接

采用同源模建方法对M1受体的三维结构进行了模拟, 将得到的模型分别与M受体完全激动剂乙酰胆碱和M1受体选择性激动剂占诺美林进行分子对接, 形成非特异性激动和特异性激动的受体-配体复合物. 用分子动力学模拟方法分别将未与小分子对接的M1受体、M1受体-乙酰胆碱复合物、M1受体-占诺美林复合物置于磷脂双膜中模拟10 ns. 将模拟后的蛋白质结构与包含活性分子的测试库对接并将结果打分, 以top5%富集因子(EF)作为评价依据, 用占诺美林优化后的M1受体模型的EF为8.0, 用乙酰胆碱优化后M1受体模型的EF为6.5, 非复合物的EF为1.5. 说明M1受体选择性激动剂复合物进行分子动力学模拟后得到的三维结构模型比较合理, 可以作为化合物虚拟筛选的模型对新化合物进行虚拟筛选, 为找到新的选择性M1受体激动剂奠定了基础.     

猪伪狂犬病毒胸苷激酶的三维结构模建与配体设计

以伪狂犬病毒(PRV)Fa为材料, 克隆测序胸苷激酶(TK)基因, 采用同源模建方法构建胸苷激酶的三维结构模型, 并经Ramachandran图和Profile_3D图验证了模型的可靠性. 采用InsightⅡ/Binding site, Delphi和Affinity方法定位了胸苷激酶的活性位点Site 1, 在此基础上设计出胸苷激酶抑制小分子N-苯基-N'-甲基脲, 通过柔性分子对接法阐明了胸苷激酶抑制剂与靶酶活性位点的相互作用模式, 发现模式中特异性的氢键相互作用可能是对靶酶产生抑制活性的重要分子基础. 研究结果为合理设计PRV胸苷激酶抑制剂, 探索新的治疗及预防伪狂犬病方案奠定了基础.     

同源模建和分子对接研究HDAC1/HDAC8的选择性

组蛋白去乙酰化酶(HDACs)是近年来治疗肿瘤的重要靶标之一.由于HDACs包含多种亚型,且各亚型的生理功能存在一定的差异,其选择性抑制剂的开发已成为当前的研发热点.我们通过同源模建的HDAC1结构,与已有的HDAC8晶体结构的活性位点进行比较分析,探讨了对两者选择性有重要影响的残基,为基于受体的选择性抑制剂研究提供重要信息.同时选择了52个HDAC抑制剂,分别建立了HDAC1、HDAC8的活性值与对接打分值的线性回归模型.所建的HDAC1和HDAC8的线性构效关系模型的非交叉验证系数R2分别为0.82和0.80,表明具有一定的统计学意义.利用所建模型对已设计合成的化合物进行了预测,预测结果对HDAC1、HDAC8选择性抑制剂的优化改造提供了一定的指导意义.     

细胞色素P450对底物的选择性及催化机制

细胞色素P450酶是广泛存在的含亚铁血红素单加氧酶,参与甾类激素的合成、脂溶性维生素代谢、多不饱和脂肪酸转换为生物活性分子,以及致癌作用和药物代谢.综述了细胞色素P450结构与功能的关系,特别是细胞色素P450对底物的选择性以及催化机制,并对其未来的发展方向进行了展望.     

细胞死亡蛋白的三维结构及活性位点的理论研究

利用同源模建和分子动力学模拟方法,构建了细胞死亡蛋白（CED-10）的三维结构,并预测了其可能的活性位点,为细胞死亡蛋白家族催化机理的深入研究提供了、重要的参考信息．     

细胞色素P450的电化学研究进展

细胞色素P450的电化学研究从一个侧面反映了为使细胞色素P450达到工业催化剂的最终目的人们所作的不懈努力。本文从细胞色素P450在电极上的电子转移研究,隧道扫描显微镜的微观成像研究和使用电极作为细胞色素P450的电子给体从而实现细胞色素P450底物转化三方面,评述了近年来细胞色素P450的电化学研究进展。     

α1A-肾上腺素受体的同源模建及其对接研究

以β2肾上腺素受体(β2-AR)为模板,采用同源模建和分子动力学模拟构建了人类α1A-肾上腺素受体(α1A-AR)的三维结构模型,并利用PROCHECK,PROSA和WHAT-IF评估了模型的合理性.所得的结构采用分子对接程序Flexidock与激动剂去甲肾上腺素和拮抗剂西罗多辛分别进行对接,结果表明,2种配基具有相似...     

Molecular characterization,modeling and docking of CYP107CB2 from Bacillus lehensis G1, an alkaliphile

Cytochrome P450s are a superfamily of heme monooxygenases which catalyze a wide range of biochemical reactions. The reactions involve the introduction of an oxygen atom into an inactivated carbon of a compound which is essential to produce an intermediate of a hydroxylated product. The diversity of chemical reactions catalyzed by cytochrome P450s has led to their increased demand in numerous industrial and biotechnology applications. A recent study showed that a gene sequence encoding a CYP was found in the genome of Bacillus lehensis G1, and this gene shared structural similarity with the bacterial vitamin D hydroxylase (Vdh) from Pseudonocardia autotrophica. The objectives of present study was to mine, for a novel CYP from a new isolate B. lehensis G1 alkaliphile and determine the biological properties and functionalities of CYP in this bacterium. Our study employed the usage of computational methods to search for the novel CYP from CYP structural databases to identify the conserved pattern, functional domain and sequence properties of the uncharacterized CYP from B. lehensis G1. A computational homology model of the protein’s structure was generated and a docking analysis was performed to provide useful structural knowledge on the enzyme’s possible substrate and their interaction. Sequence analysis indicated that the newly identified CYP, termed CYP107CB2, contained the fingerprint heme binding sequence motif FxxGxxxCxG at position 336-345 as well as other highly conserved motifs characteristic of cytochrome P450 proteins. Using docking studies, we identified Ser-79, Leu-81, Val-231, Val-279, Val-383, Ala-232, Thr-236 and Thr-283 as important active site residues capable of stabilizing interactions with several potential substrates, including vitamin D₃, 25-hydroxyvitamin D₃ and 1α-hydroxyvitamin D₃, in which all substrates docked proximally to the enzyme’s heme center. Biochemical analysis indicated that CYP107CB2 is a biologically active protein to produce 1α,25-dihydroxyvitamin D₃ from 1α-hydroxyvitamin D₃. Based on these results, we conclude that the novel CYP107CB2 identified from B. lehensis G1 is a putative vitamin D hydroxylase which is possibly capable of catalyzing the bioconversion of parental vitamin D₃ to calcitriol, or related metabolic products.     

Identification of novel Plasmodium falciparum PI4KB inhibitors as potential anti-malarial drugs: Homology modeling,molecular docking and molecular dynamics simulations

The current study was set to discover selective Plasmodium falciparum phosphatidylinositol-4-OH kinase type III beta (pfPI4KB) inhibitors as potential antimalarial agents using combined structure-based and ligand-based drug discovery approach. A comparative model of pfPI4KB was first constructed and validated using molecular docking techniques. Performance of Autodock4.2 and Vina4 software in predicting the inhibitor-PI4KB binding mode and energy was assessed based on two Test Sets: Test Set I contained five ligands with resolved crystal structures with PI4KB, while Test Set II considered eleven compounds with known IC50 value towards PI4KB. The outperformance of Autodock as compared to Vina was reported, giving a correlation coefficient (R²) value of 0.87 and 0.90 for Test Set I and Test Set II, respectively. Pharmacophore-based screening was then conducted to identify drug-like molecules from ZINC database with physicochemical similarity to two potent pfPI4KB inhibitors –namely cpa and cpb. For each query inhibitor, the best 1000 hits in terms of TanimotoCombo scores were selected and subjected to molecular docking and molecular dynamics (MD) calculations. Binding energy was then estimated using molecular mechanics–generalized Born surface area (MM-GBSA) approach over 50 ns MD simulations of the inhibitor-pfPI4KB complexes. According to the calculated MM-GBSA binding energies, ZINC78988474 and ZINC20564116 were identified as potent pfPI4KB inhibitors with binding energies better than those of cpa and cpb, with ΔG_binding ≥ −34.56 kcal/mol. The inhibitor-pfPI4KB interaction and stability were examined over 50 ns MD simulation; as well the selectivity of the identified inhibitors towards pfPI4KB over PI4KB was reported.     

Molecular docking,dynamics simulations and 3D-QSAR modeling of arylpiperazine derivatives of 3,5-dioxo-(2H,4H)-1,2,4-triazine as 5-HT1AR agonists

Serotonin receptor, 5-HT_1AR, agonists and partial agonists have established drug candidates for psychiatric and neurologic disorders. Recently, we reported the synthesis and evaluation of arylpiperazine derivatives of 3,5-dioxo-(2H,4H)-1,2,4-triazine as 5-HT_1AR ligands. Herein, we generated a homology model of the receptor and docked the ligands against it, predicted the stability of the receptor model and complexes by molecular dynamics and generated a 3D-QSAR model for the arylpiperazine derivatives of 3,5-dioxo-(2H,4H)-1,2,4-triazine. The model suggests the hydrophobic part that arises from the aromatic region and the electron withdrawing parts play a vital role in the agonist activity of the lead molecules.     

Single crystal structure of 3-(2-(3-acetylphenyl)hydrazono)pentane-2,4-dione and molecular docking study with CYP450 members for anticancer molecular screening

Coupling of acetyl acetone with diazotized 3-aminoacetophenone was carried out to give the compound 3. The compound was characterized by IR, ¹HNMR, MS and elemental analysis. The X-ray analysis of 3 revels its planar nature with torsion angles between phenyl ring and acetyl group C2–C3–C7–O1 and C2–C3–C7–C8 of 177.8(3)° and ?0.9(4)°, respectively. Small deviations from planarity are evident also by torsion angles N1–N2–C9–C10 and N2–C9–C12–O3 of 176.6(2)° and 2.9(4)°, while the N2–C9–C10–O2 torsion angle with the value of ?165.2(3)° deviates from planarity. Hydrogen-bonded chain formed through C5–H5?O2 is connected with the adjacent antiparallel chain through C11–H11B···π interaction between the methyl group of the acetyl substituent and phenyl ring forming a double-layered chain. N1–N2 shows presence of single bond, showing it to be a hydrazone. Molecular docking study of the title compound with six members from CYP450 family shows encouraging activity.     

Homology modeling and molecular dynamics study of GSK3/SHAGGY-like kinase

Although the GSK3/SHAGGY-like kinase is a highly conserved serine/threonine kinase implicated in many signaling pathways in eukaryotes, the lack of knowledge of its three-dimensional (3D) structure has hindered efforts to understand the binding specificities of substrate and catalytic mechanism. To understand the structure-activity relationships, the protein 3D structure was built by using homology modeling based on the known X-ray diffraction structure of Glycogen synthase kinase-3beta (Gsk3beta) and the model structure was further refined using unrestrained molecular dynamics simulations. The research indicates that the general 3D organization of the GSK3/SHAGGY-like kinase is a typical kinase family and comprises an N-terminal domain of beta-sheet and a larger C-terminal domain mainly constituted by alpha-helix. In order to understand the molecular interactions between the natural substrate-ATP and GSK3/SHAGGY-like kinase, a 3D model of the complex ATP-GSK3/SHAGGY-like kinase is developed by molecular docking program, which is helpful to guide the experimental realization and the new mutant designs as well. One important finding is that the identification of the key binding-site residue of Lys69 which plays an important role in the catalysis of GSK3/SHAGGY-like kinase and this is in consistent with experimental observation.     

An evaluation of molecular models of the cytochrome P450 Streptomyces griseolus enzymes P450SU1 and P450SU2

Summary P450SU1 and P450SU2 are herbicide-inducible bacterial cytochrome P450 enzymes from Streptomyces griseolus. They have two of the highest sequence identities to camphor hydroxylase (P450cam from Pseudomonas putida), the cytochrome P450 with the first known crystal structure. We have built several models of these two proteins to investigate the variability in the structures that can occur from using different modeling protocols. We looked at variability due to alignment methods, backbone loop conformations and refinement methods. We have constructed two models for each protein using two alignment algorithms, and then an additional model using an identical alignment but different loop conformations for both buried and surface loops. The alignments used to build the models were created using the Needleman-Wunsch method, adapted for multiple sequences, and a manual method that utilized both a dotmatrix search matrix and the Needleman-Wunsch method. After constructing the initial models, several energy minimization methods were used to explore the variability in the final models caused by the choice of minimization techniques. Features of cytochrome P450cam and the cytochrome P450 superfamily, such as the ferredoxin binding site, the heme binding site and the substrate binding site were used to evaluate the validity of the models. Although the final structures were very similar between the models with different alignments, active-site residues were found to be dependent on the conformations of buried loops and early stages of energy minimization. We show which regions of the active site are the most dependent on the particular methods used, and which parts of the structures seem to be independent of the methods.     

A preliminary 3D model for cytochrome P450 2D6 constructed by homology model building

Summary A homology model building study of cytochrome P450 2D6 has been carried out based on the crystal structure of cytochrome P450 101. The primary sequences of P450 101 and P450 2D6 were aligned by making use of an automated alignment procedure. This alignment was adjusted manually by matching -helices (C, D, G, I, J, K and L) and -sheets (3/4) of P450 101 that are proposed to be conserved in membrane-bound P450s (Ouzounis and Melvin [Eur. J. Biochem., 198 (1991) 307]) to the corresponding regions in the primary amino acid sequence of P450 2D6. Furthermore, -helices B, B and F were found to be conserved in P450 2D6. No significant homology between the remaining regions of P450 101 and P450 2D6 could be found and these regions were therefore deleted. A 3D model of P450 2D6 was constructed by copying the coordinates of the residues from the crystal structure of P450 101 to the corresponding residues in P450 2D6. The regions without a significant homology with P450 101 were not incorporated into the model. After energy-minimization of the resulting 3D model of P450 2D6, possible active site residues were identified by fitting the substrates debrisoquine and dextrometorphan into the proposed active site. Both substrates could be positioned into a planar pocket near the heme region formed by residues Val³⁷⁰, Pro³⁷¹, Leu³⁷², Trp³¹⁶, and part of the oxygen binding site of P450 2D6. Furthermore, the carboxylate group of either Asp¹⁰⁰ or Asp³⁰¹ was identified as a possible candidate for the proposed interaction with basic nitrogen atom(s) of the substrates. These findings are in accordance with a recently published predictive model for substrates of P450 2D6 [Koymans et al., Chem. Res. Toxicol., 5 (1992) 211].     

Molecular Dynamics Simulations of a Cytochrome P450 from Tepidiphilus thermophilus (P450-TT) Reveal How Its Substrate-Binding Channel Opens

Cytochrome P450s (P450) are important enzymes in biology with useful biochemical reactions in, for instance, drug and xenobiotics metabolisms, biotechnology, and health. Recently, the crystal structure of a new member of the CYP116B family has been resolved. This enzyme is a cytochrome P450 (CYP116B46) from Tepidiphilus thermophilus (P450-TT) and has potential for the oxy-functionalization of organic molecules such as fatty acids, terpenes, steroids, and statins. However, it was thought that the opening to its hitherto identified substrate channel was too small to allow organic molecules to enter. To investigate this, we performed molecular dynamics simulations on the enzyme. The results suggest that the crystal structure is not relaxed, possibly due to crystal packing effects, and that its tunnel structure is constrained. In addition, the simulations revealed two key amino acid residues at the mouth of the channel; a glutamyl and an arginyl. The glutamyl’s side chain tightens and relaxes the opening to the channel in conjunction with the arginyl’s, though the latter’s side chain is less dramatically changed after the initial relaxation of its conformations. Additionally, it was observed that the effect of increased temperature did not considerably affect the dynamics of the enzyme fold, including the relative solvent accessibility of the amino acid residues that make up the substrate channel wall even as compared to the changes that occurred at room temperature. Interestingly, the substrate channel became distinguishable as a prominent tunnel that is likely to accommodate small- to medium-sized organic molecules for bioconversions. That is, P450-TT has the ability to pass appropriate organic substrates to its active site through its elaborate substrate channel, and notably, is able to control or gate any molecules at the opening to this channel.