Molecular dynamics simulations of peptide carboxylate hydration

Aqueous solvation of carboxylate groups, as present in the glycine zwitterion and the dipeptide aspartylalanine, is studied employing a force-field that includes distributed multipole electrostatics and induction contributions (Amoebapro: P. Ren and J. W. Ponder, J. Comput. Chem., 2002, 23, 1497; P. Ren and J. W. Ponder, J. Phys. Chem. B, 2003, 107, 5933; J. W. Ponder and D. A. Case, Adv. Protein Chem., 2003, 66, 27). Radial and orientation distribution functions, as well as hydration numbers, are calculated and compared with existing simulation data derived from Car-Parrinello molecular dynamics (CPMD), and also distributed-charge force-fields. Connections are also made with experimental data for solvation of carboxylates in water. Our findings show that Amoebapro yields carboxylate solvation properties in very good agreement with CPMD results, significantly closer agreement than can be obtained from traditional force-fields. We also demonstrate that the influence of solvation on the conformation of the dipeptide is markedly different using Amoebapro compared with the other force-fields.     

Dynamics of water in the hydration layer of a partially unfolded structure of the protein HP-36

Atomistic molecular dynamics simulations of the folded native structure and a partially unfolded molten globule structure of the protein villin headpiece subdomain or HP-36 have been carried out with explicit solvent to explore the effects of unfolding on the dynamical behavior of water present in the hydration layers of different segments (three alpha-helices) of the protein. The calculations revealed that the unfolding of helix-2 influences the translational and rotational motions of water present in the hydration layers of the three helices in a heterogeneous manner. It is observed that a correlation exists between the unfolding of helix-2 and the microscopic kinetics of protein-water hydrogen bonds formed by its residues. This in turn has an influence on the rigidity of the hydration layers of the helices in the unfolded structure versus that in the folded native structure. These results should provide a microscopic explanation to recent solvation dynamics experiments on folded native and unfolded structures of proteins.     

Structural and dynamical analysis of the hydration of the Alzheimer's beta-amyloid peptide

An analysis of the water molecules in the first solvation shell obtained from the molecular dynamics simulation of the amyloid beta(10-35)NH2 peptide and the amyloid beta(10-35)NH2E22Q "Dutch" mutant peptide is presented. The structure, energetics, and dynamics of water in the hydration shell have been investigated using a variety of measures, including the hydrogen bond network, the water residence times for all the peptide residues, the diffusion constant, experimentally determined HN amide proton exchange, and the transition probabilities for water to move from one residue to another or into the bulk. The results of the study indicate that: (1) the water molecules at the peptide-solvent interface are organized in an ordered structure similar for the two peptide systems but different from that of the bulk, (2) the peptide structure inhibits diffusion perpendicular to the peptide surface by a factor of 3 to 5 relative to diffusion parallel to the peptide surface, which is comparable to diffusion of bulk water, (3) water in the first solvation shell shows dynamical relaxation on fast (1-2 ps) and slow (10-40 ps) time scales, (4) a novel solvent relaxation master equation is shown to capture the details of the fast relaxation of water in the peptide's first solvation shell, (5) the interaction between the peptide and the solvent is stronger in the wild type than in the E22Q mutant peptide, in agreement with earlier results obtained from computer simulations [Massi, F.; Straub, J. E. Biophys J 2001, 81, 697] correlated with the observed enhanced activity of the E22Q mutant peptide.     

Hofmeister anionic effects on hydration electric fields around water and peptide

Specific ion effects on water dynamics and local solvation structure around a peptide are important in understanding the Hofmeister series of ions and their effects on protein stability in aqueous solution. Water dynamics is essentially governed by local hydrogen-bonding interactions with surrounding water molecules producing hydration electric field on each water molecule. Here, we show that the hydration electric field on the OD bond of HOD molecule in water can be directly estimated by measuring its OD stretch infrared (IR) radiation frequency shift upon increasing ion concentration. For a variety of electrolyte solutions containing Hofmeister anions, we measured the OD stretch IR bands and estimated the hydration electric field on the OD bond to be about a hundred MV∕cm with standard deviation of tens of MV∕cm. As anion concentration increases from 1 to 6 M, the hydration electric field on the OD bond decreases by about 10%, indicating that the local H-bond network is partially broken by dissolved ions. However, the measured hydration electric fields on the OD bond and its fluctuation amplitudes for varying anions are rather independent on whether the anion is a kosmotrope or a chaotrope. To further examine the Hofmeister effects on H-bond solvation structure around a peptide bond, we examined the amide I' and II' mode frequencies of N-methylacetamide in various electrolyte D(2)O solutions. It is found that the two amide vibrational frequencies are not affected by ions, indicating that the H-bond solvation structure in the vicinity of a peptide remains the same irrespective of the concentration and character of ions. The present experimental results suggest that the Hofmeister anionic effects are not caused by direct electrostatic interactions of ions with peptide bond or water molecules in its first solvation shell. Furthermore, even though the H-bond network of water is affected by ions, thus induced change of local hydration electric field on the OD bond of HOD is not in good correlation with the well-known Hofmeister series. We anticipate that the present experimental results provide an important clue about the Hofmeister effect on protein structure and present a discussion on possible alternative mechanisms.     

Site-specific hydration status of an amphipathic peptide in AOT reverse micelles

Reverse micelles formed by sodium bis(2-ethylhexyl) sulfosuccinate (AOT) in isooctane (IO) and water have long been used as a means to provide a confined aqueous environment for various applications. In particular, AOT reverse micelles have often been used as a template to mimic membrane-water interfaces. While earlier studies have shown that membrane-binding peptides can indeed be incorporated into the polar cavity of AOT reverse micelles where they mostly fold into an alpha-helical structure, the underlying interactions leading to the ordered conformation are however not well understood. Herein, we have used circular dichroism (CD) and infrared (IR) spectroscopies in conjunction with a local IR marker (i.e., the CN group of a non-natural amino acid, p-cyano-phenylalanine) and a global IR reporter (i.e., the amide I' band of the peptide backbone) to probe the conformation as well as the hydration status of an antimicrobial peptide, mastoparan x (MPx), in AOT reverse micelles of different water contents. Our results show that at, w0=6, MPx adopts an alpha-helical conformation with both the backbone and hydrophobic side chains mostly dehydrated, whereas its backbone becomes partially hydrated at w0=20. In addition, our results suggest that the amphipathic alpha-helix so formed orients itself in such a manner that its positively charged, lysine-rich, hydrophilic face points toward the negatively charged AOT head groups, while its hydrophobic face is directed toward the polar interior of the water pool. This picture is in marked contrast to that observed for the binding of MPx to phospholipid bilayers wherein the hydrophobic surface of the bound alpha-helix is buried deeper into the membrane interior.     

Drift time mass spectrometric protein hydration experiments

Recent developments in electrospray ionization opened the field of mass spectrometry for macromolecules like proteins. The combination of ionization, drift time spectrometry, and mass spectrometry combines a large field of experimental techniques. In this new field it becomes possible to understand the interactions of isolated proteins with reaction partners like water. The combination of these techniques allows one to determine structural and thermodynamic properties. The experiments presented here give a first insight into the interaction of isolated protein ions with single solvent molecules. Results are presented for proteins like cytochrome C, bovine pancreatic trypsin inhibitor and lysozyme. MD-simulations on the same proteins and conditions as similar as possible to the experiments reflect the experimental results remarkable well.     

Bromopyridazinedione-mediated protein and peptide bioconjugation

Bromopyridazinedione-mediated bioconjugation to a cysteine containing protein and a disulfide containing peptide is described. The conjugates are cleavable in an excess of thiol, including cytoplasmically-relevant concentrations of glutathione, and show a high level of hydrolytic stability. The constructs have the potential for four points of chemical attachment.     

Preferential hydration and the exclusion of cosolvents from protein surfaces

Protein stability is enhanced by the addition of osmolytes, such as sugars and polyols and inert crowders, such as polyethylene glycols. This stability enhancement has been quantified by the preferential hydration parameter which can be determined by experiments. To understand the mechanism of protein stability enhancement, we present a statistical mechanical analysis of the preferential hydration parameter based upon Kirkwood-Buff theory. Previously, the preferential hydration parameter was interpreted in terms of the number of hydration waters, as well as the cosolvent exclusion volume. It was not clear how accurate these interpretations were, nor what the relationship is between the two. By using the Kirkwood-Buff theory and experimental data, we conclude that the contribution from the cosolvent exclusion dominantly determines the preferential hydration parameters for crowders. For osmolytes, although the cosolvent exclusion largely determines the preferential hydration parameters, the contribution from hydration may not be negligible.     

Insight into the role of hydration on protein dynamics

The potential energy surface of a protein is rough. This intrinsic energetic roughness affects diffusion, and hence the kinetics. The dynamics of a system undergoing Brownian motion on this surface in an implicit continuum solvent simulation can be tuned via the frictional drag or collision frequency to be comparable to that of experiments or explicit solvent simulations. We show that the kinetic rate constant for a local rotational isomerization in stochastic simulations with continuum solvent and a collision frequency of 2 ps(-1) is about 10(4) times faster than that in explicit water and experiments. A further increase in the collision frequency to 60 ps(-1) slows down the dynamics, but does not fully compensate for the lack of explicit water. We also show that the addition of explicit water does not only slow down the dynamics by increasing the frictional drag, but also increases the local energetic roughness of the energy landscape by as much as 1.0 kcal/mol.     

How protein surfaces induce anomalous dynamics of hydration water

Water around biomolecules slows down with respect to pure water, and both rotation and translation exhibit anomalous time dependence in the hydration shell. The origin of such behavior remains elusive. We use molecular dynamics simulations of water dynamics around several designed protein models to establish the connection between the appearance of the anomalous dynamics and water-protein interactions. For the first time we quantify the separate effect of protein topological and energetic disorder on the hydration water dynamics. When a static protein structure is simulated, we show that both types of disorder contribute to slow down water diffusion, and that allowing for protein motion, increasing the spatial dimensionality of the interface, reduces the anomalous character of hydration water. The rotation of water is, instead, altered by the energetic disorder only; indeed, when electrostatic interactions between the protein and water are switched off, water reorients even faster than in the bulk. The dynamics of water is also related to the collective structure--à voir the hydrogen bond (H-bond) network--formed by the solvent enclosing the protein surface. We show that, as expected for a full hydrated protein, when the protein surface offers pinning sites (charged or polar sites), the superficial water-water H-bond network percolates throughout the whole surface, hindering the water diffusion, whereas it does not when the protein surface lacks electrostatic interactions with water and the water diffusion is enhanced.     

Langevin model of the temperature and hydration dependence of protein vibrational dynamics

The modification of internal vibrational modes in a protein due to intraprotein anharmonicity and solvation effects is determined by performing molecular dynamics (MD) simulations of myoglobin, analyzing them using a Langevin model of the vibrational dynamics and comparing the Langevin results to a harmonic, normal mode model of the protein in vacuum. The diagonal and off-diagonal Langevin friction matrix elements, which model the roughness of the vibrational potential energy surfaces, are determined together with the vibrational potentials of mean force from the MD trajectories at 120 K and 300 K in vacuum and in solution. The frictional properties are found to be describable using simple phenomenological functions of the mode frequency, the accessible surface area, and the intraprotein interaction (the displacement vector overlap of any given mode with the other modes in the protein). The frictional damping of a vibrational mode in vacuum is found to be directly proportional to the intraprotein interaction of the mode, whereas in solution, the friction is proportional to the accessible surface area of the mode. In vacuum, the MD frequencies are lower than those of the normal modes, indicating intramolecular anharmonic broadening of the associated potential energy surfaces. Solvation has the opposite effect, increasing the large-amplitude vibrational frequencies relative to in vacuum and thus vibrationally confining the protein atoms. Frictional damping of the low-frequency modes is highly frequency dependent. In contrast to the damping effect of the solvent, the vibrational frequency increase due to solvation is relatively temperature independent, indicating that it is primarily a structural effect. The MD-derived vibrational dynamic structure factor and density of states are well reproduced by a model in which the Langevin friction and potential of mean force parameters are applied to the harmonic normal modes.     

Role of electromechanical and mechanoelectric effects in protein hydration under hydrostatic pressure

Recent measurements of lysozyme hydration water density under non-denaturing pressure show that it is higher than that of bulk water in the same conditions. High protein hydration layer density has earlier been observed at ambient conditions and ascribed to electrostriction. We calculate the pressure-induced protein mean surface charge density increment Δσ. Within the hydration layer, the higher fields due to Δσ lead to an additional water compression via electrostriction. The increment Δσ is considered as due to a mechanoelectric effect in protein molecules. The mean value of the effective mechanoelectric coefficient d is calculated and compared with piezoelectric coefficients of amino acids and their compounds.     

Fast fluctuations in protein powders: The role of hydration

An elastic neutron scattering investigation of the molecular dynamics of hydrated lysozyme powders has been undertaken for different water contents h (g water/g Lysozyme). The dry sample exhibits a harmonic behaviour in the whole temperature range, while anharmonic motions arise on hydrated samples at a temperature T_d. Both T_d and the magnitude of the anharmonic motions are markedly hydration dependent. On increasing water content the crossing barrier entropy change increases, while the enthalpy change keeps constant. The estimated average rigidity of the protein structure decreases abruptly immediately below the onset of the enzymatic activation at around 0.2h.     

Dendrimer effects on peptide and protein fibrillation

Dendrimers are synthetic, symmetrically branched polymers that can be manufactured to a high degree of definition and therefore present themselves as monodisperse entities. Flexible and globular in shape and compartementalized into a partly inaccessible interior and a highly exposed surface, they offer numerous possibilities for interactions with and responses to biological macromolecules and biostructures including cell membranes and proteins. By way of their multiple functional surface groups, they allow the design of surfaces carrying a multitude of biological motifs and/or charges giving rise to quite significant biological and physico-chemical effects. Here we describe the surprising ability of dendrimers to interact with and perturb polypeptide conformations, particularly efficiently towards amyloid structures; that is, the structures of highly insoluble polypeptide aggregates involved in a range of serious and irreversibly progressive pathological conditions (protein-misfolding diseases). Interesting as this may be, the interaction of dendrimers with such generic peptidic aggregates also offers a new perspective on the molecular mechanisms governing assembly and disassembly of amyloid structures and thereby on determinants of protein and peptide folding. Despite the potent disaggregative nature of various dendrimers, they have variable effects on the stability of different proteins, suggesting that they do not act as generic denaturants, but rather exert their effects via specific interactions with individual parts of each protein.     

Representing protein and peptide structures with parallel-coordinates

Graphical representation of molecular conformations is an important tool used by chemists to gain molecular insight. In spite of today's enhanced computer graphics there are still situations, such as in multiple conformation displays, in which standard visualization techniques are limited. Parallel-coordinate (‖-coords) representation, which was originally developed for visualizing multivariant datasets in fields other than chemistry, offers an alternative basis for graphical representation of molecular structures. In parallel-coordinates, the axes are drawn parallel rather than perpendicular to each other, allowing many axes to be placed and seen. This mapping procedure has unique geometric properties and useful relationships to the original space. In this article, we apply the parallel-coordinate representation for presenting peptide and protein structural conformations. In particular, we demonstrate the usefulness of parallel-coordinates in the context of conformational analysis where this representation, combined with multiple filters, allows nontrivial clustering of data points, leading to new observations. The ‖-coords representation is also demonstrated as a tool for two-dimensional (2D) representation of protein secondary structure and for identification of disulfide-bonded pairs in protein structures. Regardless of the application, an advantage of the ‖-coords approach is that it retains its inherent simplicity and ease of use, and requires little or no software development. © 1997 John Wiley & Sons, Inc. J Comput Chem 18 : 1893–1902, 1997     

Microencapsulation peptide and protein drugs delivery system

Many methods were used to devise peptide and protein drugs delivery system (DDS). Because of their relatively large size, they have low transdermal bioavailabilities. In systemic delivery of proteins, biodegradable material as parenteral depot formulation occupy an important place because of several aspects like protection of sensitive proteins from degradation, prolonged or modified release, pulsatile release patterns. The main objective in developing controlled release protein injectables is avoidance of regular invasive doses which in turn provide patient compliance, comfort as well as control over blood levels. This review article presents the outstanding contributions in field of microencapsulation as protein delivery systems and different approaches of protein delivery are described. Then discusses how these advances may be applied to resolve the challenges face the development of microcapsule for the controllable delivery of protein drugs.     

Far-infrared spectroscopy on free-standing protein films under defined temperature and hydration control

The functionality of proteins is governed by their dynamics. We have performed a systematic investigation on four different proteins in the far-infrared spectral region under control of the two external parameters that have the strongest influence on the dynamics, namely temperature and hydration. The absorption measurements covering the frequency range from 40 cm(-1) to 690 cm(-1) (1-20 THz) close the gap between the well-studied mid-infrared and the recent THz investigations. By preparing the proteins as free-standing films, we achieve unprecedented reproducibility. Besides a featureless slope in the THz range, we can identify absorption peaks characteristic for each protein and others common to several proteins. We fit the spectra to extract the peak positions and suggest assignments for them. The far-infrared absorption spectra of all proteins are basically independent on hydration. By a detailed analysis of the sorption isotherms this can be explained by the low absorption of biological water, which resembles more the behavior of ice than that of liquid water.     

High-resolution protein hydration NMR experiments: probing how protein surfaces interact with water and other non-covalent ligands

High-resolution solution NMR experiments are extremely useful to characterize the location and the dynamics of hydrating water molecules at atomic resolution. However, these methods are severely limited by undesired incoherent transfer pathways such as those arising from exchange-relayed intra-molecular cross-relaxation. Here, we review several complementary exchange network editing methods that can be used in conjunction with other types of NMR hydration experiments such as magnetic relaxation dispersion and ¹J_NC′ measurements to circumvent these limitations. We also review several recent contributions illustrating how the original solution hydration NMR pulse sequence architecture has inspired new approaches to map other types of non-covalent interactions going well beyond the initial scope of hydration. Specifically, we will show how hydration NMR methods have evolved and have been adapted to binding site mapping, ligand screening, protein-peptide and peptide-lipid interaction profiling.