Combining simulations and experiments for the molecular engineering of multifunctional collagen mimetic peptide-based materials.

Assembling peptides allow the creation of structurally complex materials, where amino acid selection influences resulting properties. We present a synergistic approach of experiments and simulations for examining the influence of natural and non-natural amino acid substitutions via incorporation of charged residues and a reactive handle on the thermal stability and assembly of multifunctional collagen mimetic peptides (CMPs). Experimentally, we observed inclusion of charged residues significantly decreased the melting temperature of CMP triple helices with further destabilization upon inclusion of the reactive handle. Atomistic simulations of a single CMP triple helix in explicit water showed increased residue-level and helical structural fluctuations caused by the inclusion of the reactive handle; however, these atomistic simulations cannot be used to predict changes in CMP melting transition. Coarse-grained (CG) simulations of CMPs at experimentally relevant solution conditions, showed, qualitatively, the same trends as experiments in CMP melting transition temperature with CMP design. These simulations show that when charged residues are included electrostatic repulsions significantly destabilize the CMP triple helix and that an additional inclusion of a reactive handle does not significantly change the melting transition. Based on findings from both experiments and simulations, the sequence design was refined for increased CMP triple helix thermal stability, and the reactive handle was utilized for the incorporation of the assembled CMPs within covalently crosslinked hydrogels. Overall, a unique approach was established for predicting stability of CMP triple helices for various sequences prior to synthesis, providing molecular insights for sequence design towards the creation of bulk nanostructured soft biomaterials.

Use of molecular docking computational tools in drug discovery.

Molecular docking has become an important component of the drug discovery process. Since first being developed in the 1980s, advancements in the power of computer hardware and the increasing number of and ease of access to small molecule and protein structures have contributed to the development of improved methods, making docking more popular in both industrial and academic settings. Over the years, the modalities by which docking is used to assist the different tasks of drug discovery have changed. Although initially developed and used as a standalone method, docking is now mostly employed in combination with other computational approaches within integrated workflows. Despite its invaluable contribution to the drug discovery process, molecular docking is still far from perfect. In this chapter we will provide an introduction to molecular docking and to the different docking procedures with a focus on several considerations and protocols, including protonation states, active site waters and consensus, that can greatly improve the docking results.

Role of the conformational versatility of the neurotrophin N-terminal regions in their recognition by Trk receptors.

Neurotrophins (NTs) represent a family of proteins that play an important role in the survival, development, and function of neurons. Extensive efforts are currently being made to develop small molecules endowed with agonist or antagonist NT activity. The structurally versatile N-termini of these proteins are considered regions of interest for the design of new molecules. By combining experimental and computational approaches, we analyzed the intrinsic conformational preferences of the N-termini of two of the most important NTs: NGF (NGF-Nter) and NT4 (NT4-Nter). Circular dichroism spectra clearly indicate that both peptides show a preference for random coil states. Because this finding does not preclude the possibility that structured forms may occur in solution as minor conformational states, we performed molecular-dynamics simulations to gain insights into the structural features of populated species. In line with the circular dichroism analysis, the simulations show a preference for unstructured states for both peptides. However, the simulations also show that for NT4-Nter, and to a lesser extent for NGF-Nter, helical conformations, which are required for binding to the Trk receptor, are present in the repertoire of structures that are intrinsically accessible to these peptides. Accordingly, molecular recognition of NTs by the Trk receptor is accomplished by the general mechanism known as population shift. These findings provide a structural rationale for the observed activity of synthetic peptides based on these NT regions. They also suggest strategies for the development of biologically active peptide-based compounds.

New insights and innovation from a million crystal structures in the Cambridge Structural Database.

The Cambridge Structural Database (CSD) is the world's largest and most comprehensive collection of organic, organometallic, and metal-organic crystal structure information. Analyses using the data have wide impact across the chemical sciences in allowing understanding of structural preferences. In this short review, we illustrate the more common methods by which CSD data influence molecular design. We show how more data could lead to more refined insights into the future using a simple example of trifluoromethylphenyl fragments, highlighting how with sufficient data one can build a reasonable model of geometric change in a chemical fragment with torsional rotation, and show some recent examples where the CSD has been used in conjunction with other methods to provide design ideas and more computationally tractable workflows for derivation of useful insights into structural design.

Intrinsic Structural Features of the Human IRE1α Transmembrane Domain Sense Membrane Lipid Saturation.

Activation of inositol-requiring enzyme (IRE1α) is an indispensable step in remedying the cellular stress associated with lipid perturbation in the endoplasmic reticulum (ER) membrane. IRE1α is a single-spanning ER transmembrane protein possessing both kinase and endonuclease functions, and its activation can be fully achieved through the dimerization and/or oligomerization process. How IRE1α senses membrane lipid saturation remains largely unresolved. Using both computational and experimental tools, we systematically investigated the dimerization process of the transmembrane domain (TMD) of IRE1α and found that, with help of the serine 450 residue, the conserved tryptophan 457 residue buttresses the core dimerization interface of IRE1α-TMD. BiFC (bimolecular fluorescence complementation) experiments revealed that mutation on these residues abolished the saturated fatty acid-induced dimerization in the ER membrane and subsequently inactivated IRE1α activity in vivo. Therefore, our results suggest that the structural elements of IRE1α-TMD serve as a key sensor that detects membrane aberrancy.

Hybrid Atomistic and Coarse-Grained Molecular Dynamics Simulations of Polyethylene Glycol (PEG) in Explicit Water.

In-silico design of polymeric biomaterials requires molecular dynamics (MD) simulations that retain essential atomistic/molecular details (e.g., explicit water around the biofunctional macromolecule) while simultaneously achieving large length and time scales pertinent to macroscale function. Such large-scale atomistically detailed macromolecular MD simulations with explicit solvent representation are computationally expensive. One way to overcome this limitation is to use an adaptive resolution scheme (AdResS) in which the explicit solvent molecules dynamically adopt either atomistic or coarse-grained resolution depending on their location (e.g., near or far from the macromolecule) in the system. In this study we present the feasibility and the limitations of AdResS methodology for studying polyethylene glycol (PEG) in adaptive resolution water, for varying PEG length and architecture. We first validate the AdResS methodology for such systems, by comparing PEG and solvent structure with that from all-atom simulations. We elucidate the role of the atomistic zone size and the need for calculating thermodynamic force correction within this AdResS approach to correctly reproduce the structure of PEG and water. Lastly, by varying the PEG length and architecture, we study the hydration of PEG, and the effect of PEG architectures on the structural properties of water. Changing the architecture of PEG from linear to multiarm star, we observe reduction in the solvent accessible surface area of the PEG, and an increase in the order of water molecules in the hydration shells.

Computational Design of Oligopeptide Containing Poly(ethylene glycol) Brushes for Stimuli-Responsive Drug Delivery.

Stimuli-responsive biomaterials are used to facilitate drug and gene delivery by shielding the drug/gene during circulation times and selectively releasing the cargo at the desired target. Within stimuli-responsive materials, pH-responsive materials are exploited for delivery to specific organs, intracellular compartments, cancer cells, site of inflammation or infection as those sites are characterized by pH that is different from the blood pH. In this paper we use molecular dynamics (MD) simulations to design such pH-responsive biomaterials where the balance between the various intermolecular interactions (e.g., electrostatics, van der Waals) within the biomaterials allow biofunctional molecules to be reversibly shielded and exposed to the environment with change in pH. In our model the shielding aspect is imparted by a polyethylene glycol (PEG) brush and the pH-responsive component is a PEG-tethered oligopeptide that undergoes changes in conformations via protonation of residues upon changes in pH. Starting with a PEG-tethered peptide in a monodisperse short PEG brush, we first vary the composition and sequence of histidine (H), lysine (K), and glutamate (E) along the oligopeptide sequence to find the design parameters that maximize the shielding and exposure of the oligopeptide at pH ∼ 7.0 and pH < 7.0, respectively. Then, we probe the effect of the PEG brush on the conformations of the oligopeptides by simulating PEG-tethered peptide in a bimodal PEG brush containing short PEG and long PEG chains. We characterize the intermolecular interactions involving the PEG, peptide, and solvent that influence the shielded and exposed conformations of the oligopeptides at the two different pHs. In a short monodisperse PEG brush, with a longer PEG-tethered peptide containing large blocks of histidines that undergo change in protonation state as a response to pH change, placed between a protonated lysine and deprotonated glutamate, the PEG brush exhibits maximum shielding and exposure with pH change. This change from shielded to exposed state is driven by electrostatic repulsion upon H protonation. The presence of long PEG chains in a bimodal PEG brush leads to dominating PEG-peptide attractive interactions that reduces the contrast in shielded and exposed conformations of the PEG-tethered peptide upon protonation of histidines.

Design Principles for Nanoparticles Enveloped by a Polymer-Tethered Lipid Membrane.

We propose the design for a nanoparticle carrier that combines three existing motifs into a single construct: a liposome is stabilized by anchoring it to an enclosed solid core via extended polymeric tethers that are chemically grafted to the core and physisorb into the surrounding lipid membrane. Such a design would exhibit several enticing properties, among them: (i) the anchoring stabilizes the liposome against a variety of external stresses, while preserving an aqueous compartment between core and membrane; (ii) the interplay of design parameters such as polymer length or grafting density enforces strong constraints on nanoparticle size and hence ensures a high degree of uniformity; and (iii) the physical and chemical characteristics of the individual constituents equip the construct with numerous functionalities that can be exploited in many ways. However, navigating the large parameter space requires a sound prior understanding for how various design features work together, and how this impacts potential pathways for synthesizing and assembling these nanoparticles. In this paper, we examine these connections in detail, using both soft matter theory and computer simulations at all levels of resolution. We thereby derive strong constraints on the experimentally relevant parameter space, and also propose potential equilibrium and nonequilibrium pathways for nanoparticle assembly.

Dendritic amphiphiles strongly affect the biophysical properties of DPPC bilayer membranes.

Molecular dynamics (MD) simulations were used to gain insight on the molecular interactions in a model biological membrane comprised of a bilayer with DPPC (dipalmitoylphosphotidylcholine) and antimicrobial dendritic amphiphile molecules [RCONHC(CH(2)CH(2)COOH)(3), where R is the saturated hydrocarbon tail (R = n-C(n)H(2n+1)), to be abbreviated as 3CAmn]. This study analyzes different biophysical properties of the equilibrated mixed bilayers, at 300 and 325 K, to determine how the presence of the 3CAmn, in varying concentrations and tail lengths, affects the lipid bilayer. Lipid tail order parameter data, bilayer thickness trends, and qualitative lipid tail tilt observations suggest that a molar ratio of 0.2 3CAm19/DPPC is sufficient to induce a phase transition in the bilayer from gel to liquid crystalline at 300 K. These results also imply that the phase transition temperature of the mixed bilayer decreases upon incorporation of higher concentrations of 3CAm19. Hydrogen bonding takes place between the 3CAmn and DPPC at specific sites, as evidenced by the radial distribution function. Increased hydrogen bonding and the smaller headgroup size of the 3CAmn molecule result in a decrease in the total lateral area with higher concentrations of 3CAm19. Diffusion constants of 3CAmn varied with concentration and tail length; diffusion constants of DPPC and 3CAm19 increased with increasing 3CAm19 concentration at 300 K and shorter 3CAmn tails had higher diffusion constants at both temperatures. These computational studies provide a comprehensive understanding of the biophysical changes to model biological membranes by the association of 3CAmn.

The intrinsic stability of the human prion ß-sheet region investigated by molecular dynamics.

Human prion diseases are neurodegenerative disorders associated to the misfolding of the prion protein (PrP). Common features of prion disorders are the fibrillar amyloid deposits and the formation of prefibrillar oligomeric species also suggested as the origin of cytotoxicity associated with diseases. Although the process of PrP misfolding has been extensively investigated, many crucial aspects of this process remain unclear. We have here carried out a molecular dynamics study to evaluate the intrinsic dynamics of PrP ß-sheet, a region that is believed to play a crucial role in prion aggregation. Moreover, as this region mediates protein association in dimeric assemblies frequently observed in prion crystallographic investigations, we also analyzed the dynamics of these intermolecular interactions. The extensive sampling of replica exchange shows that the native antiparallel ß-structure of the prion is endowed with a remarkable stability. Therefore, upon unfolding, the persistence of a structured ß-region may seed molecular association and influence the subsequent phases of the aggregation process. The analysis of the four-stranded ß-sheet detected in the dimeric assemblies of PrP shows a tendency of this region to form dynamical structured states. The impact on the ß-sheet structure and dynamics of disease associated point mutations has also been evaluated.

Dynamical properties of steric zipper polymorphs formed by a IAPP-derived peptide.

Understanding the molecular basis of neurodegenerative diseases has enormous implications for the development of effective therapeutic strategies. One of the most puzzling features of these pathologies is the occurrence of distinct strains, which are believed to be generated by alternative conformational transitions of the same protein/peptide. Very recently, it has been discovered that small model peptides are able to form alternative tightly packed assemblies (polymorphs) in the crystalline state. Intriguingly, it has been postulated that the different polymorphs of the same polypeptide sequence may be representative of distinct strains. As the organization of crystalline aggregates of small peptides may be heavily biased by crystal packing, we have here performed MD simulations on steric zipper polymorphs formed by of the IAPP-derived fragment SSTNVG. Our analyses show that these aggregates are rather stable also in a non-crystalline environment. This finding corroborates the hypothesis that steric zipper assemblies are good candidates to account for the phenomenon of strain in neurodegenerative diseases. Present investigations also provide clues on the factors that favour the formation of polymorphs. Indeed, the intrinsic stability of individual ß-sheets formed by SSTNVG strands is very poor. Therefore, the formation of these aggregates is essentially dictated by inter-sheet interactions established within the steric zipper assembly.

Dynamics and stability of amyloid-like steric zipper assemblies with hydrophobic dry interfaces.

Recent seminal investigations have suggested that the basic structural motif of amyloid fibers may be constituted by a tight association of two facing beta-sheets (steric zipper). Although this model has been derived from crystal structures of small peptide models, several theoretical investigations, essentially focused on steric zipper interface containing large polar and/or aromatic side chains, have confirmed the stability of this motif in a crystal-free context. To analyze the general validity of these findings, we carried out molecular dynamics (MD) simulations on aggregates stabilized by steric zipper interfaces made also of small or hydrophobic residues. In particular, we here characterized assemblies formed by the peptides SSTSAA and VQIVYK, whose structures have been recently solved at high resolution. In contrast to previous results obtained for polar/aromatic aggregates of the same size and with similar interface area, steric zipper assemblies composed of a pair of 10-stranded beta-sheets show high fluctuations and significant distortions in the simulation timescales (40-60 ns). Taking into account the crystal packing, the effect of the addition of an extra sheet to the assemblies was also evaluated. The MD results indicate that this addition does not provide extra-stabilization to the pair of sheet models. Although present data do not preclude the possibility that the steric zipper association identified in the crystal structure is the basic motif of SSTSAA and VQIVYK fibers, our findings highlight the importance of the nature of residues directly involved in the motif. Indeed, polar and aromatic residues that may form intrasheet and intersheet interactions likely provide a strong contribution to the steric zipper motif stability. Along this line, assemblies endowed with hydrophobic residues presumably require larger interfaces. In line with this suggestion, MD analysis of the HET-s(218-289) prion models composed of a similar number of strands shows that the assembly is endowed with a remarkable stability.