新型蝶啶酮类PLK1抑制剂的虚拟筛选

Polo样激酶1(PLK1)是治疗多种癌症的一个靶点,蝶啶酮衍生物作为PLK1抑制剂具有显著的生物活性。为了更好理解PLK1抑制剂的药效特征并筛选出具有新骨架的苗头化合物,对新型三/四唑蝶啶酮衍生物进行了系统的分子模拟,主要包括分子对接、药效团模型构建、虚拟筛选和分子动力学模拟。分子对接结果表明,关键氨基酸残基C67、A80、K82、L130、C133、F183可以通过疏水作用、氢键或π-π堆积作用与三/四唑蝶啶酮衍生物相互作用。通过构建的药效团模型,结合分子对接和药代动力学ADME预测,虚拟筛选得到了3个潜在化合物(VS01、VS02和VS03)。分子动力学模拟结果表明,VS02和VS03不仅能与PLK1稳定结合,而且结合自由能优于已报道活性最高的化合物27。为新型PLK1抑制剂的筛选和开发提供了有效信息和理论参考。     

葡萄糖结合蛋白（GBP）构象变化的研究及讨论

葡萄糖结合蛋白(D-glucose Binding Protein,简称GBP)属于一类在细胞膜上起化学活性物质运载相兲作用的周质结合蛋白(PBPs)。未结合葡萄糖分子的GBP (空配体GBP或"apo GBP")的构象动力学在配体结合过程中起着至兲重要的作用,了解这一过程对于设计该蛋白的抑制剂或突变具有重要意义。利用空配体GBP的全原子分子动力学模拟,确定了空配体GBP的多种亚稳态构象,模拟结果揭示了场域静电斥力或许是空配体GBP构象动力学的兲键决定因素。GBP上下两块区域的蛋白结构域相互作用,与铰链区域施加的几何约束作用相平衡。在溶液中,区域相互作用导致了空配体GBP开口构象和闭口构象乊间可以快速转化。此外,分子动力学模拟所绘制的空配体GBP的亚稳态与乊前的荧光研究一致,在核磁共振(NMR)实验中也观察到了不同构象乊间的快速平衡。结果表明,现有的晶体结构可能不代表溶液中的主要构象。     

ε-CL-20在二元混合溶剂中晶体形貌的分子动力学模拟

利用分子动力学模拟的方法探究了乙酸乙酯与二溴甲烷组成的二元溶剂在298.15 K,1 atm下对ε-CL-20晶体形貌的影响。通过修正附着能模型(MAE)模型探究了溶剂-晶体相互作用,用分子动力学模拟预测了不同组成的乙酸乙酯/二溴甲烷混合溶剂中ε-CL-20的晶体形貌并与实验获得ε-CL-20的晶体形貌进行了对比。结果表明,实验获得的晶体形貌与模拟结果一致,且晶面粗糙度越大,溶剂-晶体相互作用越强。此外,还通过均方位移(MSD)分析了溶剂分子在不同晶面的扩散系数,探究了溶剂扩散速率对不同晶面的影响,并利用径向分布函数(RDF)分析了溶剂分子与晶体分子间相互作用的组成。     

复合材料界面相互作用的分子模拟研究与进展

综述了MD(分子动力学)模拟技术、计算复合材料界面粘接力的MD模型构建与模拟方法,概括了MD在研究复合材料界面相互作用中的研究现状;最后对MD的发展方向进行了展望。     

重组B7-H1IgV融合蛋白的表达、纯化及活性鉴定

目的以B7-H1为靶点,研制肿瘤免疫治疗蛋白疫苗。方法将人B7-H1胞外片段IgV区基因插入pQE-30原核表达载体,转化大肠杆菌,经IPTG诱导表达。Ni2+-NTA亲和层析纯化蛋白,Westernblot鉴定。用纯化的rhB7-H1IgV蛋白免疫昆明小鼠,经ELISA、流式细胞术、免疫组化技术和CDC试验测定融合蛋白及其抗血清的生物学活性。结果所构建的pQE-30-TT-B7-H1IgV表达载体,能稳定表达rhB7-H1IgV蛋白,经纯化后免疫昆明小鼠,可获得高滴度抗B7-H1抗血清。经流式细胞术和免疫组化检测显示,其抗血清可与HT-29/B7-H1+及SP2/0肿瘤细胞结合,且在CDC试验中,可依赖补体杀伤HT-29/B7-H1+及SP2/0肿瘤细胞。结论rhB7-H1IgV融合蛋白不仅可引发小鼠体液免疫应答,而且其抗体还能与表达B7-H1的肿瘤细胞相结合,并介导补体依赖的体外杀伤作用。     

低聚苯酰胺蛋白质α-螺旋模拟物的合成

非肽小分子α-螺旋模拟物的设计与合成是蛋白质相互作用抑制剂研发的一个重要方向。该方法通过设计、合成有机小分子化合物来模拟蛋白质相互作用界面热点区域(hot-spot)的α-螺旋结构。利用模拟物与相应蛋白质结合,从而阻断特定蛋白质-蛋白质之间的相互作用,最终达到治疗相应疾病的目的。本文以低聚苯酰胺为骨架,设计、合成了以阿司匹林为母体的蛋白质α-螺旋模拟物。     

基于蒙特卡洛分子动力学模拟的算法在天然产物小分子与蛋白质相互作用中的应用

天然产物小分子与蛋白质相互作用的研究对于天然产物的功能研究有着非常重要的意义。作为一种快速而低成本的手段,基于分子动力学模拟的方法正在受到越来越多的重视,而基于蒙特卡洛分子动力学模拟的算法是其典型的代表。本文详细的阐述了基于蒙特卡洛分子动力学模拟的算法在天然产物小分子与蛋白质相互作用中的应用。     

天然气-水-活性剂混合体系在石英表面吸附微观机理研究

构建了天然气-水-活性剂混合体系与α-石英晶面构成的界面超分子结构模型,采用分子动力学方法研究了温度为350 K和325 K,活性剂为甲醇的情况下,岩石对天然气分子以及水分子的界面吸附行为的影响.研究发现甲醇分子可降低水分子与α-石英表面的结合能从而起到降低水锁的危害,并从分子动力学的角度解释了解除水锁危害的微观机理.     

褐煤与水微观相互作用的分子机制：多尺度分子模拟

煤与水的表面相互作用是洁净煤技术领域的重要科学问题之一。然而,褐煤与水的相互作用是一个复杂的物理化学过程,其微观机制在原子尺度和电子结构方面仍不明确。尤其缺乏褐煤中不同官能团与水之间相互作用能量、稳定性结构特征、相互作用本质的系统考察。基于多尺度分子模拟研究了褐煤与水微观相互作用的分子机制。基于量子化学计算重点考察典型褐煤模型结构中不同官能团与单分子水的相互作用,获得水分子在褐煤不同位点吸附的局域极小构象,以及稳定吸附构象的几何结构特征。基于独立梯度模型的直观绘景阐释了褐煤与水分子之间的相互作用形式主要为范德华作用和氢键。借助能量分解分析方法定量描述并确定了静电作用是稳定煤-水相互作用的最主要因素。此外,还基于分子动力学模拟揭示了不同数量褐煤分子与大量水分子相互作用的组装行为和演化特征,阐明了褐煤分子结构在大量水中的团聚现象。分子动力学模拟结果同样印证了褐煤和水分子主要以静电作用结合。     

烷烃在甲醇中无限稀释扩散的分子动力学模拟

运用分子动力学模拟方法研究了300 K和10 MPa下,正烷烃(甲烷至正癸烷)在甲醇中无限稀释扩散。模拟得到的甲醇的扩散系数与相近条件下的文献值相近,因此,进一步预测了正烷烃的扩散系数。     

Characterization of the Interaction between Gallic Acid and Lysozyme by Molecular Dynamics Simulation and Optical Spectroscopy

The binding interaction between gallic acid (GA) and lysozyme (LYS) was investigated and compared by molecular dynamics (MD) simulation and spectral techniques. The results from spectroscopy indicate that GA binds to LYS to generate a static complex. The binding constants and thermodynamic parameters were calculated. MD simulation revealed that the main driving forces for GA binding to LYS are hydrogen bonding and hydrophobic interactions. The root-mean-square deviation verified that GA and LYS bind to form a stable complex, while the root-mean-square fluctuation results showed that the stability of the GA-LYS complex at 298 K was higher than that at 310 K. The calculated free binding energies from the molecular mechanics/Poisson-Boltzmann surface area method showed that van der Waals forces and electrostatic interactions are the predominant intermolecular forces. The MD simulation was consistent with the spectral experiments. This study provides a reference for future study of the pharmacological mechanism of GA.     

Prediction of epitopes and production of monoclonal antibodies against gastric H,K-ATPase

Monoclonal antibodies (mAbs) were produced against gastric H,K-ATPase using a theoretical and experimental strategy based on prediction of linear epitopes by molecular modelling followed by production of anti- peptide antibodies. By analysing the alpha subunit sequence, we predicted several epitopes corresponding to amino acids K519-L533, E543- Y553 and S786-L798 and produced monoclonal antibodies HK519, HK543 and HK786. All three react against gastric H,K-ATPase in RaLISA, immunohistochemistry and Western blots demonstrating that they recognize the native and the SDS-denatured ionic pump and that the epitopes are located at the surface of the native ATPase. Antibody Kd are in the range 6-10x10(-8) M. Monoclonal antibody HK519 is a competitive inhibitor of ATP, in agreement with ATP binding to K519. Neither mAb 543, nor mAb 786 inhibit the ATPase activity. Monoclonal antibody 95111, whose epitope is mapped between residues C529 and E561, competes with mAb HK543 but not with the other two. We suggest that the 95111 epitope is overlapping or very close to the HK543-553 sequence. Induction of E1 conformer by binding FITC to K519 increases the number of mAb 95111 and mAb HK543 epitopes but not that of mAb 786, supporting the fact that the fragment E543-Y553 changes accessibility, maybe during the E1-E2 transconformation.      

Trastuzumab-Peptide Interactions: Mechanism and Application in Structure-Based Ligand Design

Understanding of protein-ligand interactions and its influences on protein stability is necessary in the research on all biological processes and correlative applications, for instance, the appropriate affinity ligand design for the purification of bio-drugs. In this study, computational methods were applied to identify binding site interaction details between trastuzumab and its natural receptor. Trastuzumab is an approved antibody used in the treatment of human breast cancer for patients whose tumors overexpress the HER2 (human epidermal growth factor receptor 2) protein. However, rational design of affinity ligands to keep the stability of protein during the binding process is still a challenge. Herein, molecular simulations and quantum mechanics were used on protein-ligand interaction analysis and protein ligand design. We analyzed the structure of the HER2-trastuzumab complex by molecular dynamics (MD) simulations. The interaction energies of the mutated peptides indicate that trastuzumab binds to ligand through electrostatic and hydrophobic interactions. Quantitative investigation of interactions shows that electrostatic interactions play the most important role in the binding of the peptide ligand. Prime/MM-GBSA calculations were carried out to predict the binding affinity of the designed peptide ligands. A high binding affinity and specificity peptide ligand is designed rationally with equivalent interaction energy to the wild-type octadecapeptide. The results offer new insights into affinity ligand design.     

Possible Link between Higher Transmissibility of Alpha,Kappa and Delta Variants of SARS-CoV-2 and Increased Structural Stability of Its Spike Protein and hACE2 Affinity

The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) outbreak in December 2019 has caused a global pandemic. The rapid mutation rate in the virus has created alarming situations worldwide and is being attributed to the false negativity in RT-PCR tests. It has also increased the chances of reinfection and immune escape. Recently various lineages namely, B.1.1.7 (Alpha), B.1.617.1 (Kappa), B.1.617.2 (Delta) and B.1.617.3 have caused rapid infection around the globe. To understand the biophysical perspective, we have performed molecular dynamic simulations of four different spikes (receptor binding domain)-hACE2 complexes, namely wildtype (WT), Alpha variant (N501Y spike mutant), Kappa (L452R, E484Q) and Delta (L452R, T478K), and compared their dynamics, binding energy and molecular interactions. Our results show that mutation has caused significant increase in the binding energy between the spike and hACE2 in Alpha and Kappa variants. In the case of Kappa and Delta variants, the mutations at L452R, T478K and E484Q increased the stability and intra-chain interactions in the spike protein, which may change the interaction ability of neutralizing antibodies to these spike variants. Further, we found that the Alpha variant had increased hydrogen interaction with Lys353 of hACE2 and more binding affinity in comparison to WT. The current study provides the biophysical basis for understanding the molecular mechanism and rationale behind the increase in the transmissivity and infectivity of the mutants compared to wild-type SARS-CoV-2.     

Transient Interactions of a Cytosolic Protein with Macromolecular and Vesicular Cosolutes: Unspecific and Specific Effects

Cytosolic proteins do not occur as isolated but are exposed to many interactions within a crowded cellular environment. We investigated the associations between a test cytosolic protein, human ileal bile acid binding protein (IBABP), and model cosolutes mimicking macromolecular and lipid membrane intracellular components. Using fluorescence spectroscopy, heteronuclear NMR, and molecular dynamics, we found that IBABP associated weakly with anionic lipid vesicles and experienced transient unspecific contacts with albumin. Localized dynamic perturbations were observed even in the case of apparent unspecific binding. IBABP and ubiquitin did not display mutually attractive forces, whereas IBABP associated specifically with lysozyme. A structural model of the IBABP–lysozyme complex was obtained by data‐driven docking simulation. Presumably, all the interactions shown here contribute to modulating functional communication of a protein in its native environment.     

Decoding the Molecular Effects of Atovaquone Linked Resistant Mutations on Plasmodium falciparum Cytb-ISP Complex in the Phospholipid Bilayer Membrane

Atovaquone (ATQ) is a drug used to prevent and treat malaria that functions by targeting the Plasmodium falciparum cytochrome b (PfCytb) protein. PfCytb catalyzes the transmembrane electron transfer (ET) pathway which maintains the mitochondrial membrane potential. The ubiquinol substrate binding site of the protein has heme bL, heme bH and iron-sulphur [2FE-2S] cluster cofactors that act as redox centers to aid in ET. Recent studies investigating ATQ resistance mechanisms have shown that point mutations of PfCytb confer resistance. Thus, understanding the resistance mechanisms at the molecular level via computational approaches incorporating phospholipid bilayer would help in the design of new efficacious drugs that are also capable of bypassing parasite resistance. With this knowledge gap, this article seeks to explore the effect of three drug resistant mutations Y268C, Y268N and Y268S on the PfCytb structure and function in the presence and absence of ATQ. To draw reliable conclusions, 350 ns all-atom membrane (POPC:POPE phospholipid bilayer) molecular dynamics (MD) simulations with derived metal parameters for the holo and ATQ-bound -proteins were performed. Thereafter, simulation outputs were analyzed using dynamic residue network (DRN) analysis. Across the triplicate MD runs, hydrophobic interactions, reported to be crucial in protein function were assessed. In both, the presence and absence of ATQ and a loss of key active site residue interactions were observed as a result of mutations. These active site residues included: Met 133, Trp136, Val140, Thr142, Ile258, Val259, Pro260 and Phe264. These changes to residue interactions are likely to destabilize the overall intra-protein residue communication network where the proteins’ function could be implicated. Protein dynamics of the ATQ-bound mutant complexes showed that they assumed a different pose to the wild-type, resulting in diminished residue interactions in the mutant proteins. In summary, this study presents insights on the possible effect of the mutations on ATQ drug activity causing resistance and describes accurate MD simulations in the presence of the lipid bilayer prior to conducting inhibitory drug discovery for the PfCytb-iron sulphur protein (Cytb-ISP) complex.     

The Predicted Ensemble of Low‐Energy Conformations of Human Somatostatin Receptor Subtype 5 and the Binding of Antagonists

Human somatostatin receptor subtype 5 (hSSTR5) regulates cell proliferation and hormone secretion. However, the identification of effective therapeutic small‐molecule ligands is impeded because experimental structures are not available for any SSTR subtypes. Here, we predict the ensemble of low‐energy 3D structures of hSSTR5 using a modified GPCR Ensemble of Structures in Membrane BiLayer Environment (GEnSeMBLE) complete sampling computational method. We find that this conformational ensemble displays most interhelical interactions conserved in class A G protein‐coupled receptors (GPCRs) plus seven additional interactions (e.g., Y2.43–D3.49, T3.38–S4.53, K5.64–Y3.51) likely conserved among SSTRs. We then predicted the binding sites for a series of five known antagonists, leading to predicted binding energies consistent with experimental results reported in the literature. Molecular dynamics (MD) simulation of 50 ns in explicit water and lipid retained the predicted ligand‐bound structure and formed new interaction patterns (e.g. R3.50–T6.34) consistent with the inactive μ‐opioid receptor X‐ray structure. We suggest more than six mutations for experimental validation of our prediction. The final predicted receptor conformations and antagonist binding sites provide valuable insights for designing new small‐molecule drugs targeting SSTRs.