Exploring the protein folding free energy landscape: coupling replica exchange method with P3ME/RESPA algorithm

A highly parallel replica exchange method (REM) that couples with a newly developed molecular dynamics algorithm particle-particle particle-mesh Ewald (P3ME)/RESPA has been proposed for efficient sampling of protein folding free energy landscape. The algorithm is then applied to two separate protein systems, beta-hairpin and a designed protein Trp-cage. The all-atom OPLSAA force field with an explicit solvent model is used for both protein folding simulations. Up to 64 replicas of solvated protein systems are simulated in parallel over a wide range of temperatures. The combined trajectories in temperature and configurational space allow a replica to overcome free energy barriers present at low temperatures. These large scale simulations reveal detailed results on folding mechanisms, intermediate state structures, thermodynamic properties and the temperature dependences for both protein systems.     

Simulation of coupled folding and binding of an intrinsically disordered protein in explicit solvent with metadynamics

The C-terminal domain of measles virus nucleoprotein is an intrinsically disordered protein that could bind to the X domain (XD) of phosphoprotein P to exert its physiological function. Experiments reveal that the minimal binding unit is a 21-residue α-helical molecular recognition element (α-MoRE-MeV), which adopts a fully helical conformation upon binding to XD. Due to currently limited computing power, direct simulation of this coupled folding and binding process with atomic force field in explicit solvent cannot be achieved. In this work, two advanced sampling methods, metadynamics and parallel tempering, are combined to characterize the free energy surface of this process and investigate the underlying mechanism. Starting from an unbound and partially folded state of α-MoRE-MeV, multiple folding and binding events are observed during the simulation and the energy landscape was well estimated. The results demonstrate that the isolated α-MoRE-MeV resembles a molten globule and rapidly interconverts between random coil and multiple partially helical states in solution. The coupled folding and binding process occurs through the induced fit mechanism, with the residual helical conformations providing the initial binding sites. Upon binding, α-MoRE-MeV can easily fold into helical conformation without obvious energy barriers. Two mechanisms, namely, the system tending to adopt the structure in which the free energy of isolated α-MoRE-MeV is the minimum, and the binding energy of α-MoRE-MeV to its partner protein XD tending to the minimum, jointly dominate the coupled folding and binding process. With the advanced sampling approach, more IDP systems could be simulated and common mechanisms concerning the coupled folding and binding process could be investigated in the future.     

Elucidating the catalytic mechanism of β-secretase (BACE1): A quantum mechanics/molecular mechanics (QM/MM) approach

In this quantum mechanics/molecular mechanics (QM/MM) study, the mechanisms of the hydrolytic cleavage of the Met2-Asp3 and Leu2-Asp3 peptide bonds of the amyloid precursor protein (WT-substrate) and its Swedish mutant (SW) respectively catalyzed by β-secretase (BACE1) have been investigated by explicitly including the electrostatic and steric effects of the protein environment in the calculations. BACE1 catalyzes the rate-determining step in the generation of Alzheimer amyloid beta peptides and is widely acknowledged as a promising therapeutic target. The general acid-base mechanism followed by the enzyme proceeds through the following two steps: (1) formation of the gem-diol intermediate and (2) cleavage of the peptide bond. The formation of the gem-diol intermediate occurs with the barriers of 19.6 and 16.1 kcal/mol for the WT- and SW-substrate respectively. The QM/MM energetics predict that with the barriers of 21.9 and 17.2 kcal/mol for the WT- and SW-substrate respectively the cleavage of the peptide bond occurs in the rate-determining step. The computed barriers are in excellent agreement with the measured barrier of ∼18.0 kcal/mol for the SW-substrate and in line with the experimental observation that the cleavage of this substrate is sixty times more efficient than the WT-substrate.     

Beta-hairpins with native-like and non-native hydrogen bonding patterns could form during the refolding of staphylococcal nuclease

Refolding of staphylococcal nuclease has been studied recently by hydrogen-deuterium exchange and NMR spectroscopy. These studies infer that beta-hairpin formed by strand 2 and strand 3 connected by reverse turn forms early during the refolding of nuclease. Typically, hydrogen-deuterium exchange NMR techniques are usually carried out on a time scale of milliseconds whereas beta-hairpins are known to fold on a much shorter time scale. It follows that in the experiments, the hydrogen-deuterium exchange protection patterns could be arising from a significant population of fully formed hairpins. In order to demonstrate it is the fully formed hairpins which gives rise to the hydrogen-deuterium exchange protection patterns, we have considered molecular dynamics simulation of the peptide (21)DTVKLMYKGQPMTFR(35) from staphylococcal nuclease corresponding to the beta-hairpin region, using GROMOS96 force field under NVT conditions. Starting from unfolded conformational states, the peptide folds into hairpin conformations with native-like and non-native hydrogen bonding patterns. Subsequent to folding, equilibrium conditions prevail. The computed protection factors and atom depth values, at equilibrium, of the various amide protons agree qualitatively with experimental observations. A collection of molecules following the trajectories observed in the simulations can account for experimental observations. These simulations provide a molecular picture of the formed hairpins and their conformational features during the refolding experiments on nuclease, monitored by hydrogen-deuterium exchange.     

Shielding effect in protein folding

One of the most important interactions responsible for protein folding and stability are hydrogen bonds between peptide groups. There is a constant competition between the water molecules and peptide groups in a hydrogen bond formation. Also side-chains take part in this process by reducing hydration of peptide group (shielding effect) that promotes the protein folding. In this paper, a new approach to take into account a shielding effect is presented. A modification of the energy function is derived and incorporated into the UNited RESidue (UNRES) force field. Canonical Molecular Dynamics and Replica Exchange Molecular Dynamics with UNRES force field is applied to study the influence of this effect on protein structure, folding kinetics and free energy landscapes. The results of test calculations suggest that even small contribution of this effect into energy function changes force field behavior as well as speeds up the folding process significantly.     

Modeling the alpha-helix to beta-hairpin transition mechanism and the formation of oligomeric aggregates of the fibrillogenic peptide Abeta(12-28): insights from all-atom molecular dynamics simulations

In this paper, all-atom molecular dynamics simulations in explicit solvent are used to investigate the structural and dynamical determinants of the alpha-helical to beta-hairpin conformational transition of the 12-28 fragment from the full length Abeta Alzheimer's peptide. The transition from alpha-helical to beta-structure requires the peptide to populate intermediate beta-bend geometries in which several mainly hydrophobic interactions are partially formed. This is followed by the sudden collapse to ordered beta-hairpin structures and the simultaneous disruption of the hydrophobic side-chain interactions with a consequent increase in solvent exposure. The solvent exposure of hydrophobic side-chains belonging to a sequence of five consecutive residues in the beta-hairpin defines a possible starting point for the onset of the aggregation mechanisms. Several different conformations of model oligomeric (dimeric and tetrameric) aggregates are then investigated. These simulations show that while hydrophobic contacts are important to bring together different monomers with a beta-hairpin like conformation, more specific interactions such as hydrogen-bonding and coulombic interactions, should be considered necessary to provide further stabilization and ordering to the nascent fibrillar aggregates.     

Structural and dynamical properties of different protonated states of mutant HIV-1 protease complexed with the saquinavir inhibitor studied by molecular dynamics simulations

To understand the basis of drug resistance, particularly of the HIV-1 PR, three molecular dynamics (MD) simulations of HIV-1 PR mutant species, G48V, complexed with saquinavir (SQV) in explicit aqueous solution with three protonation states, diprotonation on Asp25 and Asp25' (Di-pro) and monoprotonation on each Asp residue (Mono-25 and Mono-25'). For all three states, H-bonds between saquinavir and HIV-1 PR were formed only in the two regions, flap and active site. It was found that conformation of P2 subsite of SQV in the Mono-25 state differs substantially from the other two states. The rotation about 177 degrees from the optimal structure of the wild type was observed, the hydrogen bond between P2 and the flap residue (Val48) was broken and indirect hydrogen bonds with the three residues (Asp29, Gly27, and Asp30) were found instead. In terms of complexation energies, interaction energy of -37.3 kcal/mol for the Mono-25 state is significantly lower than those of -30.7 and -10.7kcal/mol for the Mono-25' and Di-pro states, respectively. It was found also that protonation at the Asp25 leads to a better arrangement in the catalytic dyad, i.e., the Asp25-Asp25' interaction energy of -8.8 kcal/mol of the Mono-25 is significantly lower than that of -2.6kcal/mol for the Mono-25' state. The above data suggest us to conclude that interaction in the catalytic area should be used as criteria to enhance capability in drug designing and drug screening instead of using the total inhibitor/enzyme interaction.     

Determination of protein structure and dynamics combining immune algorithms and pattern search methods

Natural proteins quickly fold into a complicated three-dimensional structure. Evolutionary algorithms have been used to predict the native structure with the lowest energy conformation of the primary sequence of a given protein. Successful structure prediction requires a free energy function sufficiently close to the true potential for the native state, as well as a method for exploring the conformational space. Protein structure prediction is a challenging problem because current potential functions have limited accuracy and the conformational space is vast. In this work, we show an innovative approach to the protein folding (PF) problem based on an hybrid Immune Algorithm (IMMALG) and a quasi-Newton method starting from a population of promising protein conformations created by the global optimizer DIRECT. The new method has been tested on Met-Enkephelin peptide, which is a paradigmatic example of multiple–minima problem, 1POLY, 1ROP and the three helix protein 1BDC. DIRECT produces an initial population of promising candidate solutions within a potentially optimal rectangle for the funnel landscape of the PF problem. Hence, IMMALG starts from a population of promising protein conformations created by the global optimizer DIRECT. The experimental results show that such a multistage approach is a competitive and effective search method in the conformational search space of real proteins, in terms of solution quality and computational cost comparing the results of the current state-of-art algorithms.     

NP-completeness of the energy barrier problem without pseudoknots and temporary arcs

Knowledge of energy barriers between pairs of secondary structures for a given DNA or RNA molecule is useful, both in understanding RNA function in biological settings and in design of programmed molecular systems. Current heuristics are not guaranteed to find the exact energy barrier, raising the question whether the energy barrier can be calculated efficiently. In this paper, we study the computational complexity of a simple formulation of the energy barrier problem, in which each base pair contributes an energy of −1 and only base pairs in the initial and final structures may be used on a folding pathway from initial to final structure. We show that this problem is NP-complete.     

氯化钾溶液中离子水化的分子动力学模拟

使用Material Studio软件包中的COMPASS力场,采用分子动力学模拟的方法研究了温度为298.15 K时,浓度分别为1.065 mol/L、2.140 mol/L、3.129 mol/L的氯化钾溶液中离子水化的微观结构和动力学性质.发现浓度对离子近程水化的结构有一定的影响,随着溶液浓度的增加O-O径向分布...     

PCA技术在二硫键连接模式预测中的应用研究

关于二硫键是由蛋白质的两个半胱氨酸之间配对形成的一种共价键,可以存在于同一条蛋白质多肽键内,也可以存在于不同的多肽键之间。二硫键的形成是蛋白质折叠过程中的重要步骤,同时影响蛋白质折叠的速率和途径。因此利用计算机方法预测二硫键连接模式有非常晕要的意义。采用一种新的方法预测二硫键连接模式。结合序列多重特征向量和通过PSIPRED得出的二级结构预测信息。由于原方法会产生高维数据,使用PCA进行降维,在降维后的低维数据上采用支持向量回归技术（SVR）来预测二硫键连接模式。结果显示,上述方法提高了预测精度。     

Investigation of the rescue mechanism catalyzed by a nucleophile mutant of rice BGlu1

In the present study, the quantum mechanical/molecular mechanical (QM/MM) method was used to investigate the rescue mechanism of an E386G mutant as well as the glycosylation mechanism of the wild rice β-d-glucosidase. E386G mutant experiences an asynchronous collaborative process to glycosylate the anionic formate with an energy barrier of 22.6 kcal/mol, while the energy barrier is 25.9 kcal/mol for the wild complex. The low energy barrier of the mutated complex suggests that anionic formate might be a good nucleophile to attack the anomeric carbon atom. Both energy barriers can be lowered when the leaving departure releases from the active site, suggesting that the product release, rather than chemistry, contributes to the rate limiting in BGlu1 mutants. Structure analyses also indicate that the external nucleophile has little steric hindrance with pocket residues and adjusts freely to proceed the rescue mechanism of the mutated complex. Our calculations provide a guide for the selectivity of exogenous nucleophiles in the future study of β-glucosidase.     

The role of loop closure propensity in the refolding of Rop protein probed by molecular dynamics simulations

Rop protein is a homo-dimer of helix-turn-helix and has relatively slow folding and unfolding rates compared to other dimeric proteins of similar size. Fluorescence studies cited in literature suggest that mutation of turn residues D30-A31 to G30-G31 (Gly₂) increases its folding and unfolding rates considerably. A further increase in number of glycines in the turn region results in decrease of folding rates compared to Gly₂ mutant. To understand the effect of glycine mutation on folding/unfolding rates of Rop and the conformational nature of turn region involved in formation of early folding species, we performed molecular dynamics simulations of turn peptides, ²⁵KLNELDADEQ³⁴ (DA peptide), ²⁵KLNELGGDEQ³⁴ (G₂ peptide), ²⁵KLNELGGGDEQ³⁵ (G₃ peptide) and ²⁵KLNELGGGEQ³⁴ (G_3′ peptide) from Rop at 300 K. Further Wt-Rop and mutant G₂-Rop monomers and dimers were also studied separately by molecular dynamics simulations. Our results show that glycine based peptides (G_n peptides) have a higher loop closure propensity compared to DA. Comparison of monomeric and dimeric Rop simulations suggests that dimeric Rop necessarily requires α_L conformation to be sampled at D30/G30 position in the turn region. Since glycine (at position 30) can readily adopt α_L conformation, G_n loop plays a dual role in both facilitating loop closure as well as facilitating reorganization/packing of helices required for structural adjustment during dimer formation in the folding of Rop. Based on our simulation results and available literature, we suggest a tentative kinetic model for Rop folding which allows us to estimate the contribution of loop closure propensity to the overall folding rates.     

基于鼠精细胞的味觉阻抗传感器 用于苦味物质检测的研究

本文设计了一种基于雄鼠精细胞的细胞阻抗传感器用于苦味物质的特异性检测.雄鼠精细胞内含有大量T2Rs受体(G蛋白偶联受体)可以对苦味物质产生特异性响应,苦味受体被苦味物质激活后产生的响应会引起细胞形态骨架的变化,从而可以被细胞阻抗传感器检测.本文探索了实验的最佳细胞密度,检测了苯硫脲和奎宁两种常见的苦味物质的响应,苯硫脲检测范围为10μmol/L~200μmol/L,检出限为4μmol/L;奎宁检测范围为62.5μmol/L~1000μmol/L,检出限为40μmol/L.传感器阻抗值增量与苦味物质浓度呈一定的线性相关性.此外,论文对该味觉传感器的特异性进行了测试分析,表明了基于细胞传感器的检测方法为代替动物和人的苦味测试提供了一种可能的新的手段.     

Identification of phenoxyacetamide derivatives as novel DOT1L inhibitors via docking screening and molecular dynamics simulation

Dot1-like protein (DOT1L) is a histone methyltransferase that has become a novel and promising target for acute leukemias bearing mixed lineage leukemia (MLL) gene rearrangements. In this study, a hierarchical docking-based virtual screening combined with molecular dynamic (MD) simulation was performed to identify DOT1L inhibitors with novel scaffolds. Consequently, 8 top-ranked hits were eventually identified and were further subjected to MD simulation. It was indicated that all hits could reach equilibrium with DOT1L in the MD simulation and further binding free energy calculations suggested that phenoxyacetamide-derived hits such as L01, L03, L04 and L05 exhibited remarkably higher binding affinity compared to other hits. Among them, L03 showed both the lowest glide score (−12.281) and the most favorable binding free energy (−303.9 +/− 16.5 kJ/mol), thereby making it a promising lead for further optimization.     

一种分布式网格计算框架以及在大规模分子动力学模拟中的应用

分子动力学（molecular dynamics）模拟蛋白质等大分子内原子间的相互作用,蛋白质折叠所需的时间通常在微秒（10^-6s）量级，而进行模拟的时间步长在飞秒（10^-15s）量级，并且每步需要计算大量的相互作用（O（n^2），n为原子数），以致于无法模拟足够长时间的折叠过程．现今在满足精确度的需求下没有更好的模拟算法．最近，生物学家研究了一种分布式的动力学方法，使得可以利用分布在Internet上的计算机进行并行模批成为可能,本文的目标是设计并实现在分布式P2P和网格计算环境等多种异构计算资源下进行动力学模拟的可靠框架，以便更大限度地利用计算资源，加快计算过程．我们基于Java和web service技术，已经实现了对应用透明的计算框架，并已将它扩展到我们的网格计算环境,实验表明分子动力学模拟程序在该框架下运行良好．     

类弹性蛋白多肽的分子动力学研究

类弹性蛋白多肽因其具有特殊的相变性质,故而在重组蛋白纯化方面展现出良好的应用前景,对其发生相变的机理进行研究具有重要意义.本文利用同源建模的方法构建了类弹性蛋白多肽的三维结构并进行能量优化,之后采用分子动力学模拟手段,在300 K～400 K间5个不同温度下,对含有100个氨基酸残基的类弹性蛋白多肽[KV8F-20]各进行了6 ns的模拟.模拟过程中,类弹性蛋白多肽发生疏水缩聚,初始结构变得更加紧凑,且温度越高折叠程度越大.水分子在类弹性蛋白多肽的相变行为中起到关键作用.经分析,结果发现疏水作用与水的排出在类弹性蛋白多肽发生相变过程中起到关键作用,类弹性蛋白多肽随着温度的升高,在疏水作用驱动下,其构象折叠程度因疏水缩聚而变得更大.此外,比较不同温度下蛋白的缩聚程度,推断类弹性蛋白多肽在375 K左右发生相变,这与实验观测的结果基本吻合.据此,推测该类弹性蛋白多肽变温度范围约为95℃～102℃.这对后续调控类弹性蛋白多肽的相变具有指导作用.     

Exploring the potential of complex formation between a mutant DNA and the wild type protein counterpart: a MM and MD simulation approach

We have demonstrated that the methods of molecular modeling and molecular dynamics simulation might be used to assess whether a specific mutation in the DNA would destabilize a known DNA-protein complex. The approach is based on probing the changes in the interaction that would be induced into the complex if within the already formed wild type complex the mutation could be introduced. We have used Hoxc8-DNA complex as a test system where it is known that the Hoxc8 binding affinity of the DNA is completely lost upon mutation of the DNA by replacing TAAT stretch to GCCG. Mutation was obtained by changing the relevant base pairs into the DNA of the model of the corresponding wild type complex developed by homology modeling and MD simulation in water for 2.0 ns. Comparison of the structure, dynamics and interactions between the hypothetical mutant model with those of the similarly refined wild type model shows that the loss of affinity of the mutant DNA to Hoxc8 has two different origins: (i) loss of several strong H-bonds as the direct consequences of mutation and (ii) reduced H-bonds in the common parts due to a net loss or inferior H-bonding geometry induced by the mutation as indirect effects. The net change in the interaction energy between the DNA and the protein in the best possible configuration indicated the experimentally observed destabilization effects. No significant change in the groove width was observed and no correlation was found between the water-bridges and the loss of affinity.     

改进二进制量子粒子群算法在蛋白质折叠中的应用*

为了改善已有二维HP模型蛋白质折叠算法容易陷入局部最优、找不到理论最低能量构象的缺点,提出一种基于变异算子的改进二进制量子粒子群算法。采用二进制编码蛋白质序列,提出变异策略,并采用惩罚因子避免出现蛋白质重叠,最后将该算法应用于蛋白质序列进行测试。测试结果表明,改进算法能够找到更优的结果,算法具有一定的实用性和有效性。