Molecular Dynamics Simulations Of Biological Macromolecules

生物大分子的分子动力学模拟

• 批准号: 8939759
• 负责人: Bernard R Brooks
• 金额: $ 49.06万
• 依托单位: NATIONAL HEART, LUNG, AND BLOOD INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8939759
• 关键词: ATP Hydrolysis; Address; Affect; Affinity; Agitation; Amino Acids; Atlases; Basal Ganglia; Basic Science; Behavior; Binding; Bioinformatics; Biological; Biophysics; C-terminal; Ca(2+)-Transporting ATPase; Calcium ion; Cell physiology; Cells; Cereals; Collaborations; Complex; Computational Biology; Computing Methodologies; Cyclic AMP; Cyclic Nucleotides; Cytoplasm; Data Set; Development; Disease; Electron Microscopy; Electrons; Enzymes; Event; Family; Filtration; Frequencies; Galactosidase; Gene Expression Microarray Analysis; Goals; Human Genetics; Image Analysis; Integral Membrane Protein; Ion Channel; Kidney Diseases; Kidney Failure; Kinesin; Laboratories; Ligands; Measurement; Meiosis; Membrane; Methods; Microscopic; Microtubules; Mitosis; Modeling; Molecular; Molecular Biology; Molecular Conformation; Molecular Models; Motion; Motor; Movement; Muscle; Muscle Cells; Mutate; N Domain; NPHS2 protein; National Heart, Lung, and Blood Institute; Nerve; Nucleotides; Pathway interactions; Pharmaceutical Preparations; Phosphorylation; Phosphorylation Site; Play; Process; Protein Dephosphorylation; Proteins; Proteinuria; Protons; Publishing; Quantum Mechanics; Research; Research Project Grants; Resolution; Reticulum; Role; SERCA1; Sampling; Sarcoplasmic Reticulum; Science; Scientist; Self Perception; Signal Transduction; Sinoatrial Node; Site; Solutions; Spinal Ganglia; Structure; Structure-Activity Relationship; Synapses; System; Techniques; Testing; Threonine; Time; Trefoil Motif; United States National Institutes of Health; Walking; Water; Work; X-Ray Crystallography; base; biophysical properties; computer studies; conformational conversion; design; dimer; evaluation/testing; in vivo; insight; interest; macromolecule; models and simulation; molecular dynamics; molecular mechanics; molecular modeling; novel; podocyte; polarized cell; polypeptide; programs; simulation; slit diaphragm; small molecule; tandem mass spectrometry; theories; tool

Several diverse projects are being pursued. These are the major ones pursued during the past year. Coarse-grained and all-atom molecular dynamics of an ion channel Hyperpolarization-activated cyclic nucleotide-gated 2 (HCN2) ion channels are expressed in the sinoatrial node, dorsal root ganglia and the basal ganglia. They play fundamental roles in electric signaling in nerve, muscle and synapse, but their function and gating mechanism are not completely understood. The overall goal of the project is to gain insight into the mechanism of the HCN2 channel activation upon binding of cyclic adenosine monophosphate (cAMP) to its intracellular C-terminal. Many mechanisms have been proposed for the opening motion propagation in the channel, but they do not completely explain the entire channel behavior. A novel theory states that upon cAMP binding, a part of the HCN2 C-terminal, called the C-helix, stabilizes its secondary structure and moves towards the binding pocket to make contacts with cAMP. Its movement is correlated with the opening conformational change of the channel pore. This theory is being tested using a novel computational method, the self-guided Langevin dynamics (SGLD), which employs guided forces to enhance the low-frequency motion and accelerate the protein conformational search. Starting from the holo state structure and using the distances from tmFRET measurements as constrains, the protein is guided into its apo state. The simulations enable sampling of conformations along this transition, giving insight into the occurring structural changes and ultimately into the HCN2 gating mechanism. Coarse-grained simulations of HCN2 bring information about the folding pathway and additional insight into the stability of the protein with and without ligand. Computational study of the β-galactosidase This work is a collaboration with a cryo-EM laboratory within the NIH. It was sparked by their publishing of a solution structure at 3.2 resolution of β-galactosidase, one of the most used enzymes in molecular biology. Its long sequence (1024 aminoacids) and the fact that it forms a tetramer in order to function, make it an ideal system to apply SGLD and Coarse-grained modeling. This study will add robustness to the cryo-EM technique by comparing the structure and dynamics of the solution cryo-EM structure and the previously published X-ray crystallography structures. Calcium ATPase Conformational Transition through Self-Guided Langevin Dynamics Simulation The sarcoplasmic reticulum (SR) Ca2+-ATPase (SERCA1a) transport calcium ions from cytoplasm into the reticulum and relaxes the muscle cells. Many crystal structures of SERCA1 in various binding states have been determined, which provide insights into the mechanism of transport Ca2+ across the membrane. Molecular modeling and simulation studies are also devoted to the understanding of this important process. SERCA1a is an integral membrane protein. It comprises a single polypeptide chain of 994 amino acid residues. It is clear from the crystal structures that SERCA has a 10 helices trans-membrane domain (M), an actuator domain (A), a nucleotide binding domain (N), and a phosphorylation domain (P). The Ca2+ transport cycle starts with Ca2E1 through the Ca2+ dependent phosphorylation by ATP, leading to the formation of the Ca2E1P high-energy intermediate. Ca2E1P transits to Ca2E2P, which releases Ca2+ into the lumen of SR and leads to the E2P state. After dephosphorylation, E2P transits to E2 state and closes the luminal gate. Through thermo agitation, E2 transits to E1 by releasing protons into the cytoplasm. E1 has high Ca2+ affinity and binds with Ca2+ to form Ca2E1. To understand the transport mechanism, it is desirable to study the dynamic process during the conformation transition. Self-guided Langevin dynamics (SGLD) is a simulation method capable of studying events with large conformational change. SGLD simulations of SERCA at different binding states produce conformational transitions between conformational states. New conformations for E1.2Ca2+ and E2.P state have been identified and at E2 state the crystal structure is a preferred conformation. Atomic mechanism of the kinesin walking on microtubule Kinesin is a protein belonging to the class of Cytoskeletal motor proteins. Kinesin converts the energy of ATP hydrolysis into stepping movement along microtubules, which supports several vital cellular functions including mitosis, meiosis, and the transport of cellular cargo. Because kinesin is a fundamental protein, further research on the topic will provide important information as to how it functions. Combined with low resolution electron microscopic images, self-guided Langevin dynamics simulations are performed to study molecular motion and conformational change of kinesin motor domain in water and binding with microtubule. SGLD enable simulation to reach the time scale required for conformational change to understand the role of ATP binding and interaction with microtubules. Analysis of the glomerular phosphoproteome Diseases of the kidney filtration barrier are a leading cause of endstage renal failure. Most disorders affect the podocytes, polarized cells that are connected by a unique cell junctional complex, the slit diaphragm. Podocytes require tightly controlled signaling to maintain their integrity, viability and function. Here we provide an atlas of in vivo phosphorylated, glomerulus-expressed proteins including podocyte-specific gene products identified in an unbiased tandem mass spectrometry-based approach. We discovered 2,449 phosphorylated proteins corresponding to 4,171 identified high-confident phosphorylated residues and performed a systematic bioinformatics analysis of this dataset. Among the 146 phosphorylation sites found on proteins abundantly expressed in podocytes, several sites resided close to residues known to be mutated in human genetic forms of proteinuria. One such site discovered on the slit diaphragm protein Podocin, threonine-234 (T234), resides at the interface of Podocin dimers with a distance between both T234 residues of less than 10 Angstrom. We show that phosphorylation critically regulates dimer formation and that this may represent a general principle for the assembly of the large family of PHB-domain containing proteins.

几个不同的项目正在进行中。这些是过去一年中所追求的主要目标。 离子通道的粗粒度和全原子分子动力学 超极化激活的环核苷酸门控 2 (HCN2) 离子通道在窦房结、背根神经节和基底神经节中表达。它们在神经、肌肉和突触的电信号传导中发挥着重要作用，但它们的功能和门控机制尚不完全清楚。该项目的总体目标是深入了解环磷酸腺苷 (cAMP) 与其胞内 C 末端结合后 HCN2 通道激活的机制。对于通道中的开放运动传播，人们提出了许多机制，但它们并没有完全解释整个通道的行为。一种新颖的理论指出，在 cAMP 结合后，HCN2 C 末端的一部分（称为 C 螺旋）会稳定其二级结构，并向结合袋移动以与 cAMP 接触。其运动与通道孔的开放构象变化相关。该理论正在使用一种新的计算方法——自引导朗之万动力学（SGLD）进行测试，该方法利用引导力来增强低频运动并加速蛋白质构象搜索。从全息态结构开始，并使用 tmFRET 测量的距离作为约束，将蛋白质引导至其 apo 态。模拟能够对这一转变过程中的构象进行采样，从而深入了解发生的结构变化并最终了解 HCN2 门控机制。 HCN2 的粗粒度模拟带来了有关折叠途径的信息以及对有或没有配体的蛋白质稳定性的额外见解。 β-半乳糖苷酶的计算研究 这项工作是与 NIH 内的冷冻电镜实验室合作进行的。这是由他们发表的 β-半乳糖苷酶（分子生物学中最常用的酶之一）分辨率为 3.2 的溶液结构引发的。它的长序列（1024 个氨基酸）以及为了发挥功能而形成四聚体的事实，使其成为应用 SGLD 和粗粒度建模的理想系统。这项研究将通过比较溶液冷冻电镜结构和之前发表的 X 射线晶体学结构的结构和动力学，增加冷冻电镜技术的稳健性。 通过自引导朗之万动力学模拟实现钙 ATP 酶构象转变 肌浆网 (SR) Ca2+-ATP 酶 (SERCA1a) 将钙离子从细胞质转运到网织并放松肌肉细胞。 SERCA1 在各种结合状态下的许多晶体结构已被确定，这为了解 Ca2+ 跨膜转运机制提供了见解。 分子建模和模拟研究也致力于理解这一重要过程。 SERCA1a 是一种整合膜蛋白。 它包含 994 个氨基酸残基的单条多肽链。 从晶体结构可以清楚地看出，SERCA具有10个螺旋的跨膜结构域(M)、致动结构域(A)、核苷酸结合结构域(N)和磷酸化结构域(P)。 Ca2+ 运输循环从 Ca2E1 开始，通过 ATP 进行 Ca2+ 依赖性磷酸化，从而形成 Ca2E1P 高能中间体。 Ca2E1P转变为Ca2E2P，将Ca2+释放到SR管腔中并导致E2P状态。去磷酸化后，E2P 转变为 E2 态并关闭管腔门。 通过热搅拌，E2 通过将质子释放到细胞质中转变为 E1。 E1具有高Ca2+亲和力，与Ca2+结合形成Ca2E1。 为了了解运输机制，需要研究构象转变期间的动态过程。 自引导朗之万动力学（SGLD）是一种能够研究具有较大构象变化的事件的模拟方法。 SERCA 在不同结合状态下的 SGLD 模拟会产生构象状态之间的构象转变。 E1.2Ca2+ 和 E2.P 态的新构象已被识别，并且在 E2 态晶体结构是优选的构象。 驱动蛋白在微管上行走的原子机制 驱动蛋白是属于细胞骨架运动蛋白类的蛋白质。驱动蛋白将 ATP 水解的能量转化为沿着微管的步进运动，支持多种重要的细胞功能，包括有丝分裂、减数分裂和细胞货物运输。由于驱动蛋白是一种基本蛋白质，因此对该主题的进一步研究将提供有关其功能的重要信息。 结合低分辨率电子显微镜图像，进行自引导朗之万动力学模拟，研究水中驱动蛋白运动域的分子运动和构象变化以及与微管的结合。 SGLD 使模拟能够达到构象变化所需的时间尺度，以了解 ATP 结合以及与微管相互作用的作用。 肾小球磷酸蛋白质组分析 肾脏滤过屏障疾病是终末期肾衰竭的主要原因。大多数疾病都会影响足细胞，即通过独特的细胞连接复合体（狭缝隔膜）连接的极化细胞。足细胞需要严格控制的信号传导以维持其完整性、活力和功能。在这里，我们提供了体内磷酸化、肾小球表达的蛋白质的图谱，包括通过无偏串联质谱法鉴定的足细胞特异性基因产物。我们发现了 2,449 个磷酸化蛋白质，对应于 4,171 个已识别的高可信度磷酸化残基，并对这个数据集进行了系统的生物信息学分析。在足细胞中大量表达的蛋白质上发现的 146 个磷酸化位点中，有几个位点靠近已知在人类蛋白尿遗传形式中发生突变的残基。在狭缝隔膜蛋白 Podocin 上发现的一个这样的位点，苏氨酸-234 (T234)，位于 Podocin 二聚体的界面处，两个 T234 残基之间的距离小于 10 埃。我们发现磷酸化关键性地调节二聚体的形成，这可能代表了含有 PHB 结构域的蛋白质大家族组装的一般原则。