Exploring the conformational dynamics of alanine dipeptide in solution subjected to an external electric field: A nonequilibrium molecular dynamics simulation

In this paper, we investigate the conformational dynamics of alanine dipeptide under an external electric field by nonequilibrium molecular dynamics simulation. We consider the case of a constant and of an oscillatory field. In this context we propose a procedure to implement the temperature control, which removes the irrelevant thermal effects of the field. For the constant field different time-scales are identified in the conformational, dipole moment, and orientational dynamics. Moreover, we prove that the solvent structure only marginally changes when the external field is switched on. In the case of oscillatory field, the conformational changes are shown to be as strong as in the previous case, and non-trivial nonequilibrium circular paths in the conformation space are revealed by calculating the integrated net probability fluxes.Comment: 23 pages, 12 figure

Atomistic insights into the effects of electromagnetic fields on an amyloidogenic peptide

In this thesis computational modelling was used to investigate on the atomistic scale, the effects of external electric and electromagnetic fields (EMF) on protein behaviour and responses in an aqueous environment. The motivation behind this study is discussed in Chapter 1, along with a literature review on previous research on the effects of EMF on proteins, from both simulation based studies and experimental research. Non-Equilibrium Molecular Dynamics was employed to simulate the protein response to external perturbations and the computational methodologies encompassing this technique are described in Chapter 2. The history of NEMD is also described in this chapter, along with the specific computational details for the simulation of our peptide model, apoC-II(60-70) exposed to EMFs of static/oscillating nature and different frequency and strength. In Chapter we used all-atom NEMD simulations to understand and quantify the response mechanisms of the amyloidogenic apoC-II (60-70) peptides to non-ionising radiation by modelling their behaviour under static electromagnetic and electric fields ofdifferent strengths. Chapter 4 expands on the previous study by investigating the role of the field frequency on the peptide behaviour in solution. Finally, Chapter 5 presents a summary of the conclusions drawn from this work

Conformational equilibria and spectroscopy of gas-phase homologous peptides from first principles

Peptides and proteins fulfil crucial tasks enabling and maintaining life. Their function is directly correlated with their three-dimensional structure, which is in turn determined by their chemical composition, the amino-acid sequence. Predicting the structure of a peptide based only on its sequence information is of fundamental interest. A fully first-principles treatment free of empirical parameters would be ideal. However, this presents an ongoing challenge, due to the large system size and conformational space of most peptides. In the present work, we address this challenge concentrating on the example of polyalanine-based peptides in the gas phase. Such studies under isolated conditions follow a bottom-up approach that allows one to investigate the intramolecular interactions important for secondary structure separate from environmental effects. Furthermore, direct benchmarks of theoretical structure predictions against experiment are facilitated. The peptide series Ac-Alan-Lys(H+), (n > 6), forms α-helices in the gas phase due to a favorable interaction of the helix dipole with the positive charge at the C-terminal lysine residue. Using this design principle as a template, we explore the impact of increased structural flexibility on the conformational space due to (i) sequence length [Ac-Alan-Lys(H+), n = 19], (ii) charge placement [Ac-Ala19-Lys(H+) versus Ac-Lys(H+)-Ala19], and (iii) backbone elongation of the monomer units as represented by β-amino acids [Ac-β2hAla6-Lys(H+)]. To address the large conformational space, we develop a three-step structure-search strategy employing an unprecedented first-principles screening effort. After pre-sampling of the conformational space using a force field, thousands of structures are optimized employing density-functional theory (DFT). For this, the PBE functional is used, coupled with a pairwise correction for van der Waals interactions. For the best few structure candidates, ab initio replica-exchange molecular-dynamics simulations are performed in order to refine the local structural environment. It is shown that these can yield lower-energy conformations and lead to rearrangements of the hydrogen-bonding network. In order to connect to experiment, collision cross sections are calculated that link to ion mobility-mass spectrometry. Furthermore, infrared spectra are derived from ab initio Born-Oppenheimer molecular-dynamics simulations accounting for anharmonicities within the classical-nuclei approximation. As expected, the 20-residue peptide Ac-Ala19-Lys(H+) forms helical structures. In contrast, placing the charge at the N-terminus [Ac-Lys(H+)-Ala19], leads to several different compact structures, which are close in energy. Such small energy differences present a challenge to the theoretical approach. Incorporating exact exchange and many-body van der Waals effects predicts the presence of only one dominant conformer, which is compatible with both experimental datasets. In comparison to Ac-Ala6-Lys(H+), the β-peptide Ac-β2hAla6-Lys(H+) exhibits increased conformational flexibility due to an extended monomer backbone. Out of the almost 15,000 structures optimized with DFT, no helical conformers are found in the low-energy regime. This is changed when considering vibrational free energy (300K, harmonic approximation), which strongly favors helical conformations due to softer vibrational modes. One possible structure candidate is the H16-helix, which is compatible with both experiments. It is a unique structure as it exhibits a hydrogen-bonding pattern equivalent to the helix of natural peptides. The systems considered here highlight the advances of current DFT functionals to address the large conformational space of peptides, but also the need for further development

A computational study of cyclic peptides with vibrational circular dichroism

Cyclic peptides are a class of molecules that has shown antimicrobial potential. These are complex compounds to investigate with their large conformational space and multiple chiral centers. A technique that can be used to investigate both conformational preferences and absolute configuration (AC) is vibrational circular dichroism (VCD). To extract information from the experimental VCD spectra a comparison with calculated spectra is often needed and this is the focus of this thesis: the calculation of VCD spectra. The VCD spectra are very sensitive to small structural changes, and to accurately calculate the spectra, all important conformers need to be identified. The first part of this thesis has been to establish a reliable computational protocol using meta-dynamics to sample the conformational space and ab initio methods to calculate the spectra for cyclic peptides. Using our protocol, we have investigated if VCD alone can determine the AC of cyclic tetra- and hexapeptides. We show that it is possible to determine the AC of the cyclic peptides with two chiral centers while for the peptides with three and four chiral centers, VCD is at best able to reduce the number of possible ACs and further investigation with other techniques is needed. Further, we investigated four cyclic hexapeptides with antimicrobial potential. These peptides, in contrast to the ones used for validating the protocol, consist of several amino acids with long and positively charged side chains. For these peptides, a molecular dynamics based approach provided VCD spectra in better agreement with experiment than our protocol. Reasons for this may be the lack of atomistic detail in the solvent model used during the conformational search and insufficient description of dispersion interactions during the meta-dynamics simulation

Towards an understanding of peptide-inorganic interactions

This study has contributed to the developing understanding of fundamental principles through which interactions at peptide-inorganic interfaces occur. The inorganic materials; crystalline zinc oxide (ZnO) and platinum (Pt) together with specific binding peptides identified using the phage display technique and alanine scanning for mutant sequences selected on the basis of peptide stability calculated in silico were synthesized, extensively characterized and the mechanisms of their interaction and the effects thereof studied. Firstly ZnO growth was monitored during solution synthesis from precursors using two different ZnO methods in the absence and presence of ZnO binding peptides (ZnO-BPs); G-12 (GLHVMHKVAPPR), its mutants (G-12A6, G-12A11, G-12A12) and GT-16 (GLHVMHKVAPPR-GGGC). Secondly, adsorption characteristics and thermodynamics of interaction of ZnO with ZnO-BPs and Pt with platinum binding peptides (Pt-BPs) were studied using biophysical tools; quartz crystal microbalance with dissipation monitoring (QCM-D) and isothermal titration calorimetry (ITC)

Interplay between chirality and dynamics of complex systems: a novel computational approach

The aim of this PhD project was to set-up a computational procedure for accurate and economic Electronic Circular Dichroism (ECD) calculations of complex systems, such as biomolecules or nanostructures. This goal was achieved combining classical molecular dynamics (MD) simulations, essential dynamics (ED) analysis, and state-of-the-art time-dependent density functional theory (TDDFT) calculations. The procedure was tested on several classes of systems, switching from small peptides in water to gold nanoclusters soluble in different solvent environments. By means of this protocol, we were able to extract the most probable conformers, explicitly include the solvent, and obtain a final statistically averaged ECD for each system under investigation. In all the cases, a good qualitative agreement between the experimental and calculated ECD spectra was obtained, thus confirming the reliability of the procedure. It is worth noting that, for the first time, conformational effects and explicit water molecules have been included in the ECD calculation of thiolate-protected gold nanoclusters. The method was also used to investigate the direct role of the aqueous solvent on chiroptical properties and vice versa, showing that the water itself can assume a chiral arrangement due to the solute-solvent interactions. Moreover, the scheme was employed to study the correlation between the conformational landscape in solution and the solid-state evolution of heterochiral Phe-based dipeptides. Therefore, this affordable, yet accurate, scheme for the computation of ECD spectra has shown its ability to reproduce different experimental situations, giving insight into the chiroptical properties of complex systems.The aim of this PhD project was to set-up a computational procedure for accurate and economic Electronic Circular Dichroism (ECD) calculations of complex systems, such as biomolecules or nanostructures. This goal was achieved combining classical molecular dynamics (MD) simulations, essential dynamics (ED) analysis, and state-of-the-art time-dependent density functional theory (TDDFT) calculations. The procedure was tested on several classes of systems, switching from small peptides in water to gold nanoclusters soluble in different solvent environments. By means of this protocol, we were able to extract the most probable conformers, explicitly include the solvent, and obtain a final statistically averaged ECD for each system under investigation. In all the cases, a good qualitative agreement between the experimental and calculated ECD spectra was obtained, thus confirming the reliability of the procedure. It is worth noting that, for the first time, conformational effects and explicit water molecules have been included in the ECD calculation of thiolate-protected gold nanoclusters. The method was also used to investigate the direct role of the aqueous solvent on chiroptical properties and vice versa, showing that the water itself can assume a chiral arrangement due to the solute-solvent interactions. Moreover, the scheme was employed to study the correlation between the conformational landscape in solution and the solid-state evolution of heterochiral Phe-based dipeptides. Therefore, this affordable, yet accurate, scheme for the computation of ECD spectra has shown its ability to reproduce different experimental situations, giving insight into the chiroptical properties of complex systems

Structural and Conformational Analysis of B-cell Epitopes − component to guide peptide vaccine design

Peptide vaccines have many potential advantages including low cost, lack of need for cold-chain storage and safety. However, it is well known that approximately 90% of B-cell Epitopes (BCEs) are discontinuous in nature making it difficult to mimic them for creating vaccines. To perform a detailed structural analysis of these epitopes, they needs to be mapped onto antigen structures that are complexed with antibody. In order to obtain a clean dataset of antibody-antigen complex crystal structures, a pipeline was designed to process automatically and clean the antibody related structures from the PDB. To store this processed antibody structural data, a database (AbDb) was built and made available online. The degree of discontinuity in B-cell epitopes and their conformational nature was studied by mapping epitopes in the antibody-antigen dataset. The discontinuity of B-cell epitopes was analysed by defining extended ‘regions’ (R, consisting of at least 3 antibody-contacting residues each separated by ≤ 3 residues) and small fragments (F, antibody-contacting residues that do not satisfy the requirements for a region). Secondly, an algorithm was developed to classify region shape as linear, curved or folded. Molecular dynamics simulations were carried out on isolated epitope regions (wild type and mutant peptides). The mutant peptides have been designed by mutating non-contacting and hydrophobic residues in epitopes. Two types of mutations (hy- drophobic to alanine and hydrophobic to glutamine) have been studied using molec- ular dynamics simulations. Furthermore, the effect of end-capping on wild type and mutant epitope regions has been studied. Simulation studies were carried out on 5 linear and 5 folded shape regions. Out of these, 2 epitopes (one linear and one folded), along with their mutants and derivatives, were tested experimentally for conformational stability by CD spectroscopy and NMR. The binding of isolated epitopes with antibody was also validated by ELISA and SPR

Structural Investigation of the Complex of Filamin A Repeat 21 with Integrin αIIb and β3 Cytoplasmic Tails – A Potential “Transmission” to Regulate Cell Migration

Cell functions in multi-cellular organisms are strongly depend on the dynamic cooperation between cell adhesion and cytoskeleton reorganization. Integrins, the major cell adhesion receptors, bind to extracellular matrix (ECM) and soluble ligands on the cell surface and link to the actin cytoskeleton inside the cell membrane. In this manner, integrins integrate cell adhesion and cytoskeleton reorganization by acting as a mechanical force transducer and a biochemical signaling hub (Zamir and Geiger 2001). Consequently, integrins are vital for development, immune responses, leukocyte traffic and hemostasis, and a variety of other cellular and physiological processes. Integrins are also are the focal point of many human diseases, including genetic, autoimmune, cardiovascular and others. In terms of the cell-ECM adhesion, integrins can exist in two major states, active, where it binds to appropriate extracellular ligands, and inactive, where it disassociates from extracellular ligands. The cellular pathways that modify the integrin extracellular ligand binding states have been called inside-out integrin signaling while the pathways that are mediated by the extracellular binding have been called outside-in integrin signaling. Although the directions of outside-in and inside-out signaling point to each other, they often happen reciprocally. Rather than just integrins alone accomplishing integrin signaling, numerous proteins are recruited around integrins and are limited to the clearly defined range of focal adhesion that are large molecular complexes containing \u3e100 proteins which link integrins to cytoskeleton (Figure 1) (Zaidel-Bar et al. 2004). Proteins that directly interact with integrins are crucial for understanding integrin signaling. More importantly, proteins that link integrins to the cytoskeleton are responsible for both mechanical forces and biochemical signal transduction, as well as reorganizing the cytoskeleton. Moreover, the modification of integrin ligand binding states is dependent on the linkage t