Enhanced Stability of the Model Mini-protein in Amino Acid Ionic Liquids and Their Aqueous Solutions

Using molecular dynamics simulations, the structure of model mini-protein was thoroughly characterized in the imidazolium-based amino acid ionic liquids and their aqueous solutions. We report that the mini-protein is more stable when AAIL is added as a cosolvent. Complete substitution of water by organic cations and anions further results in hindered conformational flexibility of the mini-protein. This observation suggests that AAILs are able to defend proteins from thermally induced denaturation. We show by means of radial distributions that the mini-protein is efficiently solvated by both solvents due to agood mutual miscibility. However, amino acid based anions prevail in the first coordination sphere of the mini-protein

Methods Development and Force Field Evaluation for Molecular Simulations of Iinteractions Between Structured Peptides and Functionalized Material Surfaces

The process of protein adsorption to material surfaces is highly complex and it is one of the most fundamental concepts upon which progress in the field of bioengineering is based. The strategic design of material surfaces for optimal utility in specific biological environments is absolutely dependent upon a thorough understanding of the mechanisms underlying protein adsorption, yet there is still a very limited understanding of these mechanisms. The primary reason for this lack of understanding is that protein adsorption is a dynamic process which occurs at the atomic and macromolecular scale, where experimental analyses provide a view that is static and too coarse to elucidate the stepwise processes behind this critical biochemical phenomenon. In recent years, continual improvements in speed and efficiency of computational hardware and simulation techniques have enabled the use of molecular simulation for studying systems of the size necessary for examining the mechanistic details of protein adsorption (tens to hundreds of thousands of atoms). Of the various forms of molecular simulation, all-atom empirical force field molecular dynamics (MD) simulation has shown the greatest potential for exploring the nature of protein adsorption because it offers a dynamic view of nanosecond-scale processes with atomistic detail. However, a shortcoming of the application of MD in studying protein adsorption is that the most widely used MD force fields (i.e., equations and parameter sets used for calculating structural and energetic properties) have been designed and validated for simulations of solvated molecular systems in the absence of solid surfaces. To address this shortcoming of an otherwise extremely powerful research tool, an initial evaluation of the applicability of existing MD force fields to model systems of structured peptides interacting with functionalized material surfaces is warranted. The work presented here encompasses that initial evaluation of force fields. Numerous detailed analyses of water, ions, and peptides were completed in order to provide the most accurate and comprehensive examination of simulated peptide adsorption available. As a result of this work, simulation methods for these unique systems were tested and determined to be appropriate for accurately representing experimental results. Also, a comparative evaluation of force field performance identified the force field that most consistently reflects experimental findings

DEVELOPMENT OF EXPERIMENTAL METHODS TO ASSESS ADSORPTION FREE ENERGY FOR PEPTIDE-SURFACE INTERACTIONS

Because of its governing role in cellular response to implants and substrates for biomedical applications, the understanding and control of protein adsorption to material surfaces has been one of the major topics of research in the field of biomaterials. Unfortunately, it has proven to be extremely difficult to quantitatively understand and control these types of interactions because of the complexities involved, and existing methods that have been developed and used to characterize protein-surface interactions have proved to be inadequate to provide the level of detail necessary to achieve this understanding. New, more fundamental methods, both experimental and computational, are needed to overcome the current limitations. At a fundamental level, protein adsorption behavior can be considered to be represented by the combination of the individual interactions between the amino acid residues making up a protein, the solvent environment, and the functional groups presented by a surface. These interactions can be best characterized by the standard state adsorption free energy associated with their adsorption to a functionalized surface, and this information could be potentially very useful for understanding the sub-molecular events that govern protein adsorption behavior. In this dissertation, we specifically develop experimental methods for the determination of free energy to quantitatively characterize peptide adsorption behavior to well-defined surfaces presenting functional groups common to many types of polymeric biomaterials using surface plasmon resonance (SPR) spectroscopy. Also, because SPR is primarily limited to the types of surfaces that can readily be formed as thin layers in nanometer scale on gold biosensor substrates, methods are further developed and applied to enable values of free energy to be determined for peptide adsorption to any microscopically flat surface. The development and application of these methods enables the fundamental aspects underlying protein adsorption behavior to be characterized and provides data that can be used for the evaluation, modification, and validation of computational models that may be used to accurately predict protein adsorption behavior

Potential and Free Energy Surfaces of Adsorbed Peptides

Peptide adsorption on solid surfaces is a common process that occurs in nanotechnology and biology, with applications in the formation of nanomaterials, biosensing and drug delivery, amongst many others. Peptide adsorption involves complex processes that are difficult to characterise experimentally. Computational approaches such as molecular dynamics (MD) are often employed to better understand biomolecular systems. However, the computationally demanding nature of such systems combined with the long characteristic timescales of peptide adsorption means MD is not well suited to its study with current computing capacities. An alternative computational approach to characterising the behaviour of atoms and molecules is mapping the potential energy surface (PES) – the molecular energy as a function of the positions of all atoms – by determining its local energy minima and saddle points, which represent stable configurations and transition states that lie between them. These minima and saddle points may be located using optimisation algorithms. Harmonic approximations yield information about transition rates between minima via saddle points as well as the free energy surface (FES). This methodology – which is referred to as ‘energy landscape mapping’ (ELM) hereafter – is able to characterise fast and slow processes equally, only being limited by the size and complexity of the system studied, and the applicability of the potential energy models used. In the past, it has largely been applied to atomic and molecular clusters, and to biomolecules. It has never been applied to adsorption of peptides or any other biomolecule. In three journal papers included in this thesis, this approach is for the first time applied to adsorbed peptides. Firstly, ELM was applied to polyalanine adsorbed on surfaces of varying interactions strengths. Results obtained were comparable results to those obtained in a prior study of the same system using an evolutionary algorithm. In the second paper, ELM was applied to met-enkephalin at a gas/graphite interface, and compared with a molecular simulation technique designed for accelerating the simulation of slow processes, replica exchange molecular dynamics (REMD). In the final paper, ELM was applied to two met-enkephalin molecules at a gas/graphite interface, introducing an additional level of complexity and a step towards practical application, given real peptide adsorption processes often occur en masse. In all of these studies, information about transitions between conformations, energy barriers, rates, and the nature of the overall PES and FES, all of which were previously unknown for the systems studied, was obtained by ELM. The work conducted here has demonstrated the applicability of ELM to peptide/surface systems. Future work may consist of applying ELM to other similar processes of practical importance, developing and validating potential energy models suitable for modelling interfacial systems, including the effect of solvents, and continual development of the methodology to accelerate calculations.Thesis (Ph.D.) -- University of Adelaide, School of Chemical Engineering and Advanced Materials, 202