Role of Arginine in Mediating Protein–Carbon Nanotube Interactions

Article pubs.acs.org/Langmuir

Role of Arginine in Mediating Protein−Carbon Nanotube Interactions Eugene Wu,† Marc-Olivier Coppens,‡ and Shekhar Garde*,† †

Howard P. Isermann Department of Chemical and Biological Engineering and Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York 12180, United States ‡ Department of Chemical Engineering, University College London, Torrington Place, London, WC1E 7JE, United Kingdom S Supporting Information *

ABSTRACT: Arginine-rich proteins (e.g., lysozyme) or poly-Larginine peptides have been suggested as solvating and dispersing agents for single-wall carbon nanotubes (CNTs) in water. In addition, protein structure−function in porous and hydrophobic materials is of broad interest. The amino acid residue, arginine (Arg+), has been implicated as an important mediator of protein/ peptide−CNT interactions. To understand the structural and thermodynamic aspects of this interaction at the molecular level, we employ molecular dynamics (MD) simulations of the protein lysozyme in the interior of a CNT, as well as of free solutions of Arg+ in the presence of a CNT. To dissect the Arg+−CNT interaction further, we also perform simulations of aqueous solutions of the guanidinium ion (Gdm+) and the norvaline (Nva) residue in the presence of a CNT. We show that the interactions of lysozyme with the CNT are mediated by the surface Arg+ residues. The strong interaction of Arg+ residue with the CNT is primarily driven by the favorable interactions of the Gdm+ group with the CNT wall. The Gdm+ group is not as well-hydrated on its flat sides, which binds to the CNT wall. This is consistent with a similar binding of Gdm+ ions to a hydrophobic polymer. In contrast, the Nva residue, which lacks the Gdm+ group, binds to the CNT weakly. We present details of the free energy of binding, molecular structure, and dynamics of these solutes on the CNT surface. Our results highlight the important role of Arg+ residues in protein−CNT or protein-carbon-based material interactions. Such interactions could be manipulated precisely through protein engineering, thereby offering control over protein orientation and structure on CNTs, graphene, or other hydrophobic interfaces.

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for binding to and splicing RNA,21,22 and shuttling between the nucleus and the cytoplasm.23 The TAT protein in HIV-1 and other cell-penetrating peptides are known to be argininerich.24−26 Lysozyme and histones, also arginine-rich proteins, have been used to disperse CNTs in water.13,27 Single-strained DNA has also been used to disperse and separate CNTs of different chiralities.28−31 Recent experiments and molecular dynamics (MD) simulations show that poly-L-arginine can disperse CNTs in water.16 Free arginine (Arg+) is commonly used as an additive to stabilize therapeutic protein formulations, especially at high protein concentrations, to prevent protein aggregation.32−34 Arginine has also been shown to interact favorably with caffeic acid35 and other aromatic organic solutes and drug molecules in aqueous solutions, increasing their solubility.36,37 Finally, arginine (one of only three amino acids, with the other two being tryptophan and cysteine) has been shown to induce gelation of graphene oxide,38 suggesting favorable interactions and possibly binding of arginine to graphene oxide.

INTRODUCTION Interactions of proteins with carbon-based materials (buckyballs, carbon nanotubes, graphene, and others) are of interest in many applications, such as biosensors,1,2 nanotoxicology,3 and in the development of hybrid biomaterials.4 Surfaces of folded globular proteins are complex, and display a mixture of polar or charged residues and some hydrophobic residues to the aqueous environment. Several studies have shown that folded proteins/peptides can bind to solid hydrophobic interfaces, such as carbon nanotubes5−9 and inside of hydrophobic pores,10 and can remain functional in the bound state.4,10−12 Because interfaces of most unfunctionalized carbon-based materials are hydrophobic, one expects individual hydrophobic/aromatic residues or hydrophobic patches on protein surfaces to interact strongly with such materials.13,14 Interestingly, recent experimental and computational studies15−17 (as well as the simulations described below) suggest that arginine residues on protein surfaces, which are positively charged, may also play an important role in the binding/anchoring of proteins to CNT or graphene surfaces. On average, ∼5% of the total amino acids of a typical protein is arginine,18 which is almost always surface-exposed.19,20 Arginine-rich regions in the SR family of proteins are known © XXXX American Chemical Society

Received: November 5, 2014 Revised: January 8, 2015

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Langmuir In the above studies, the guanidinium group (Gdm+) of arginine is implicated in the interactions of arginine with carbon-based materials or with aromatic compounds in water.16,17,35,36 Specifically, the less favorable (hydrophobiclike) hydration and π−π interactions of the planar faces of the Gdm+ moiety are thought to contribute favorably to those interactions.16,17 Here, we employ MD simulations to study the details of arginine−carbon nanotube interactions in aqueous solutions. To quantify the contributions of the Gdm+ group and the remaining part of arginine (norvaline), we perform simulations of carbon nanotubes in solutions of arginine, as well as guanidinium hydrochloride and norvaline. Guanidinium chloride is a protein denaturant39 at high concentrations and has been suggested to induce dehydration of CNTs40 and unfold hydrophobic polymers.41 While the biochemistry and interactions of norvaline are interesting in other contexts,42−44 its use here serves to dissect the contributions of two different parts of arginine−CNT interactions.

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Figure 1. (A) Snapshot of the initial configuration of lysozyme (O-1 orientation as seen in panel (B)) in a 43,43 CNT (cyan, licorice). A portion of the CNT and some water molecules were removed to show the confined lysozyme. Cl− ions are shown in gray, space-fill. The remaining water molecules are shown as licorice (red, oxygen; white, hydrogen). The box is 12 nm × 12 nm × 12 nm. (B) Illustrations of four different starting orientations of lysozyme studied (top), with the rotated axis (relative to the axis in O-1) shown beneath each snapshot.

METHODS

We performed two types of simulations, one containing the protein lysozyme (Protein DataBank code 1AKI) and a carbon nanotube in water, and another containing a CNT solvated in aqueous solutions of arginine chloride, guanidinium chloride, or norvaline molecules. We describe each of the components and systems below. Carbon Nanotubes. The chirality and diameter of CNTs are determined by the indices n and m. All CNTs we simulated are armchair single-walled CNTs, for which n = m, all built using the Nanotube Builder VMD.45 The CNT was frozen at the center of the simulation box. The atom type CA in the AMBER force field was used to describe the interactions of carbons of the CNTs, with LennardJones C−C parameters of σ = 0.34 nm and ϵ = 0.36 kJ/mol, and a partial charge of zero. That is, we do not use a polarizable force field or a quantum treatment of electrons to represent the aromaticity of the CNT. In this sense, we follow others who represent the carbons of the CNT using a classical description for studies of solvation, wetability, and interactions.46−49 Because the CNT is frozen, intratube C−C interactions are excluded, since they have no effect of the properties studied here. Simulations of Lysozyme in a CNT. Our system includes a single lysozyme molecule placed at the center of a 10-nm-long (43,43), single-walled CNT with an inside diameter of ∼6 nm. As noted above, the carbon atoms of the CNT are fixed to their locations. Thus, the CNT simulates a pristine cylindrical hydrophobic pore in which to study a protein. We understand that a very small piece of a singlewalled CNT with such a large diameter is known to collapse;50,51 nevertheless, the results on binding of the protein, lysozyme mediated by arginine (as shown below), are robust and applicable to proteins in or near hydrophobic interfaces. Lysozyme was represented explicitly at the atomic level using the AMBER99SB force field.52 Eight Cl− ions are added to the solution to neutralize the net charge on lysozyme. We performed four separate simulations with four different relative orientations of lysozyme placed inside the CNT to study the nature of protein−CNT binding. A representative snapshot is shown in Figure 1A. Of these, one of the simulations (Figure 1B, O-1) was performed in a larger box (approximately 12 nm × 12 nm × 12 nm) containing 53 811 water molecules, and the other three (Figure 1B, O2−O-4) with 28 639 water molecules (approximately 12 nm × 9 nm × 9 nm). Since we are only interested in the protein confined inside of the CNT, we reduced the size of the box by decreasing the number of water molecules on the outside of the CNT to minimize computational expense in the latter three simulations. A 1-ns canonical ensemble (NVT) equilibrium simulation was performed, followed by a 1-ns isothermal−isobaric ensemble (NPT) equilibration prior to the production run. Production runs of 10 ns for the larger system and 5 ns for the three smaller systems were performed in the NPT ensemble. Other simulation details are described below.

CNTs in Aqueous Solutions of Arginine (Arg+), Guanidinium (Gdm+), and Norvaline (Nva). Systems used in solution simulations included a single (n,n) single-walled CNT, 4000 water molecules, and either 10 arginine chloride pairs, 10 guanidinium chloride pairs, or 10 norvaline (neutral) molecules. A representative snapshot of the guanidinium chloride−CNT−water system is shown in Figure 2D. We used (n,n) CNTs of five different indices (n = 6, 9, 12, 15, and 18, having diameters of d ≈ 0.8, 1.2, 1.6, 2.0, and 2.4 nm, respectively) and a length of 2.5 nm in these simulations. For guanidinium chloride and norvaline solutions, only one concentration (∼0.13 M (mol/L), corresponding to 10 ion pairs or 10 Nva solutes) was studied, whereas for arginine chloride systems, we also sampled concentrations of 0.13,

Figure 2. Space-fill representations and chemical structures of free (A) arginine (Arg+), (B) norvaline (Nva), and (C) guanidinium ion (Gdm+). Panel (D) shows a snapshot of the starting configuration for a dilute Gdm+ (cyan, carbon; blue, nitrogen; and white, hydrogen; space-fill) system with the CNT (cyan, licorice) in the center of the box along with Cl− counterions (gray, space-fill) and water molecules (red, oxygen; white, hydrogen; licorice). B

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Figure 3. (A) Lysozyme center of mass (COM) trajectory (dark blue to dark red for 0−5 ns, respectively) shows binding in the radial direction. Circle shown is 3 nm in diameter for reference. (B) Snapshots at 5 ns (gray, transparent represent lysozyme; cyan, licorice represent CNT) with atoms in Arg within 4 Å of CNT (space-fill; green, Gdm+ group; blue, Nva group). (C) Lysozyme can move in the axial direction of the CNT when bound. (D) O-1, O-2, and O-4 bind to the CNT within 2 ns while O-3 starts bound and moves toward the surface. [Note that the color scheme for panels (C) and (D) are given in panel (A).] (E) Absolute difference between Arg (R) and its average radial COM for O-1. See Figure 1 for starting orientations. 0.25, 0.47, 0.87, 1.23, 1.55, and 1.83 M solvating each CNT. Each concentration was achieved by varying the number of Arg+, Cl−, and water molecules. All molecules were placed randomly within the simulation box and all systems were electrically neutral overall. All molecules were represented explicitly in atomic detail. For arginine chloride and norvaline, we used the AMBER94 force field,53 whereas, for guanidinium chloride ions, we used the OPLS force field54 as employed by other researchers.39,41 The representation of Gdm+ groups in Arg+ and in Gdm+ are very similar to each other. For the Arg+ and Nva, the force field parameters were taken from the Nterminal (NTER) and C-terminal (CTER) parameters of the arginine residue in AMBER94. The original charges for each atom in the molecule were modified to achieve a net molecule charge of +1 for Arg+ and 0 for Nva (see the Supporting Information for details). Parameters for Molecular Dynamics (MD) Simulations. All simulations were performed using the MD package GROMACS,55 with water molecules represented explicitly using the TIP3P model.54 The leapfrog algorithm was used with a time step of 2 fs to integrate the equations of motion. The temperature was maintained at 300 K with a Nosé−Hoover thermostat,56 while the pressure was maintained at 1 bar with a Parrinello−Rahman barostat.57 Equilibration runs in each simulation included a 1-ns simulation in NVT ensemble followed by a 1-ns simulation in the NPT ensemble. Finally, in each solution system, a 20-ns NPT ensemble production run was performed with frames stored every 1 ps for further analysis. The average box size in solution simulations was approximately 5 nm × 5 nm × 5 nm. Periodic boundary conditions were used in all simulations. The particle mesh Ewald (PME) algorithm was used in all simulations to calculate electrostatic interactions58 with a grid spacing of 0.12 nm and a cutoff of 1.0 nm. A 1.0 nm cutoff was also applied to the Lennard-Jones interactions. In the simulations of lysozyme inside a CNT, a slightly

longer cutoff of 1.3 nm was used both for Lennard-Jones interactions and for PME. Lorentz−Berthelot mixing rules were used for cross Lennard-Jones interactions.59 Bonds containing hydrogen atoms were constrained using the P-LINCS algorithm.60 Standard deviations were calculated using blocks of 4 ns. Umbrella Sampling Simulations. We employed the umbrella sampling method to obtain the potential mean force (PMF) between solutes−the arginine ion (Arg+), the guanidinium ion (Gdm+), and norvaline (Nva)−and a (6,6) CNT along the radial direction at the center of the CNT. The umbrella simulations probed only the region outside of the (6,6) nanotube. For Nva molecules, the umbrella potential was applied on the center of their Cα atom, whereas for the Gdm+ ion, it was applied on the central carbon atom. Since the arginine ion contains both a Cα and guanidinium group with a central carbon atom, the umbrella potential was applied to each of these atoms in separate umbrella sampling simulations. As described previously, the CNT was held fixed at the center of the simulation box. Each umbrella window included one CNT and one solute molecule solvated by ∼4000 water molecules. The umbrella potential was harmonic, 0.5K(r − r0)2, where K is the spring constant, r the instantaneous location of the atom on which the umbrella potential is applied, and r0 the reference location of that atom (i.e., the center of the umbrella) in that window. The separations (center of CNT to center of Cα) in arginine and norvaline CNT systems were sampled using 11 windows placed at r0 = 0.86, 1.0, 1.1, 1.25, 1.375, 1.5, 1.625, 1.75, and 2.0 nm. For the Arg+ ion and Gdm+ ion CNT PMF calculations, eight windows were used, with r0 = 0.8, 0.9, 1.0, 1.2, 1.4, 1.6, 1.8, and 2.0 nm from center of the CNT to the central carbon atom of the guanidinium group or guanidinium ion, respectively. All simulations used a value of K = 500 (kJ/mol)/nm2. For each window, the simulation was run for 10 ns and frames were stored every 0.5 ps C

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Figure 4. (A) Lysozyme COM trajectory for O-1 with r̃COM (where r̃COM = rCOM − ⟨rCOM⟩) shows binding within 2.5 ns. Five snapshots are labeled and shown below. (B) For O-4, lysozyme shows binding within 2 ns. Four snapshots along the trajectory are labeled and shown below. When any atom of the residue was within 4 Å of the CNT wall, arginine was colored red while all others were colored cyan, both in space-fill. The protein is shown in gray, and the CNT is shown in cyan licorice. See Figure 1 for starting orientations. for analysis. Other simulation parameters were identical to that described above. The PMF was calculated using the weighted histogram analysis method (WHAM).61 The first 500 ps were considered as an equilibration step. From 0.5−9.5 ns, standard deviations were calculated using blocks of 1.9 ns.

the orientation of the protein in the bound state (shown in gray in Figure 3B) is influenced by the initial configuration, as expected. Figure 3B illustrates the guanidinium group of arginine (green) interacting with the interior surface, with the majority lying parallel to the surface. In the original starting configuration (O-1), once arginine residues come into contact with the surface, they remain bound for the remainder of the simulation, as seen in Figure 3E. However, what is most interesting is the dynamics of attachment of the protein to the CNT surface. In all simulations, arginine residues play an important role in establishing firm contact of the protein with the CNT wall and helping to anchor the protein. For example, in trajectory O1, R128 as well as Proline70 (P70) make fleeting contacts with the CNT over a time scale of 0.5 ns. However, it is the R128 of lysozyme that makes firm contact with the CNT surface, anchoring the protein, followed by P70, R68, R21, and other contacts, as shown in the time series of pictures in Figure 4. A similar picture with arginine residues helping the protein establish first contact of the protein with the CNT also emerges from visualization of other trajectories (see Figure 4 for trajectory O-4). The importance of arginine in protein−CNT interactions has also been highlighted recently by other researchers. For example, in simulations of lysozyme on the outside of a (10,10) CNT, Calvaresi et al. identified seven residues that were primary contributors to the free energy of binding (ΔGbinding > 4 kcal/mol ≈ 16.7 kJ/mol). Of those seven, two are arginines (R21 and R112) located on two different regions: one in the 310-helix and the other in the α-

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RESULTS AND DISCUSSION Role of Arginine in Lysozyme−CNT Interactions. Our simulations of the protein, lysozyme, inside a CNT were motivated by experimental data on its activity when confined inside ordered mesoporous silica having different pore sizes.10 The stability, folding, and enzymatic activity of proteins and the behavior of water under confinement are different under confinement, compared to that in bulk solution. The effects of confinement on both protein and water have been studied extensively.10,46,47,62−69 In our simulations, we noticed that arginine residues (denoted as R) on the surface of lysozyme helped anchor it to the CNT wall. We include relevant results here simply to highlight this point before proceeding to a moredetailed study of arginine−CNT interactions. Figure 3 summarizes the results of MD simulations of lysozyme in the interior of a (43,43) armchair CNT. We performed four separate simulations each beginning with a different orientation of lysozyme at the CNT center. Figure 3A shows that, in all four simulations, lysozyme diffuses toward the CNT wall, binds the CNT within ∼1−2 ns, and remains bound during the remainder of the simulation. In the bound state, the protein moves along the wall in the angular and axial directions, as seen clearly in Figures 3A, 3C, and 3D. In the confined simulations, D

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Favorable water-mediated interactions of all three molecules with the CNT are reflected in the peaks near the surface of the CNTs (Figure 5). We have plotted the absolute densities, and we have not normalized them by the bulk density. As these systems are relatively dilute solutions, the bulk density is small, and upon normalization, the details of these distributions get washed out due to the very large peaks. Of the three solutes, Gdm+ appears to have the most favorable interactions, followed by Arg+, whereas Nva displays a weaker affinity for the CNT. Arg+ and Nva have more-complex geometries, which result in different orientations on the surface, leading to the shoulders or twin-peaked density distributions, especially for Nva on the inside of the (18,18) CNT. The smaller normalization volume inside the (18,18) CNT also amplifies these features, as observed in Figure 5B. The relative locations of the first peaks suggest that Gdm+ COM is able to get closest to the CNT, whereas the Nva peak is farther out. Figure 6 shows snapshots from three simulations,

helix D. Calvaresi et al. stated that these two structural motifs bind the CNT as a tweezer by using their arginine residues (R21 and R112), which provide the most important contribution to the overall binding.15 Interactions of R128 with CNT are involved in the binding of lysozyme to CNT in all four trajectories studied here. R128 is present in a flexible loop adjacent to Leu-129 at the C-terminus and is fully solvent exposed. Kubiak-Ossowska and Mulheran also identified R128 as a major contributor to protein-charged-surface and protein− protein interactions of lysozyme,70 and showed that mutations at R128 weaken surface adsorption.71 These simulations of lysozyme inside a CNT serve to highlight the role of surface arginine residues in protein−CNT interactions. Below, we quantify, at the molecular detail, the structural, dynamic, and thermodynamic aspects of watermediated arginine−CNT interactions.

Figure 6. Snapshots of a (6,6) CNT in (A) Arg+Cl−, (B) Gdm+Cl−, and (C) Nva solutions. Arg+, Gdm+, Nva, Cl−, and the CNT are shown in a space-fill representation. Water molecules are shown (red, oxygen; white, hydrogen; licorice). The bottom of each panel focuses on the vicinity of the CNT to show different modes of solute−CNT binding. Water molecules are removed from these snapshots for the sake of clarity (blue, nitrogen; white, hydrogen; red, oxygen; and cyan, carbon). Figure 5. Equilibrium COM density profiles of Arg+, Gdm+, and Nva plotted as a function of r* (where r* = rCOM − rCNT) near (A) (6,6) CNT and (B) (18,18) CNT. The vertical dashed line at r* = 0 indicates the CNT surface (i.e., carbon atom centers): for r* > 0, a molecule is on the outside, and for r* < 0, a molecule is on the inside.

which highlight the local aggregation of Arg+ and Gdm+ near the CNT, and the weaker interactions of Nva with the CNT. Figure 6B shows that Gdm+ is a flat, disk-like molecule and orients itself parallel to the surface of the CNT, consistent with the strong first peak in the density profile. Previous work by Godawat et al.41 has also shown that the Gdm+ ions coat a hydrophobic polymer and unfold it, with surface parallel orientations dominating the interactions. The Arg+ molecules make several contacts with the CNT, with the dominant contact being that of the Gdm+ group of Arg+ lying flat on the surface of the CNT. This qualitative picture suggests that Arg+−CNT interactions are likely driven by the interactions of the Gdm+ group of Arg+, with other moieties of Arg+ making small additional contributions. In the absence of the Gdm+ group, Nva molecules interact weakly with the CNT. Nva− CNT contacts involve interactions of the aliphatic side chain of Nva with CNT. In the presence of the charged carboxyl and amino groups in the vicinity, these interactions are not as strong as the typical hydrophobic interactions. We also

Density Distribution of Arg+, Gdm+, and Nva near CNTs. Figure 5 shows equilibrium center of mass (COM) density profiles in the radial direction for Arg+, Gdm+, and Nva molecules near (6,6) and (18,18) CNTs obtained from simulations in 0.13 M aqueous solutions of these solutes. (See Supporting Information for intermediate CNT sizes and different CNT parameters.) To make comparisons between distributions near CNTs of different sizes, the profiles are plotted as a function of r* = rCOM − rCNT, which is the COM distance from the CNT surface. To focus on the watermediated interactions between CNT and these solutes, only molecules having their COM within the body portion of the CNT were considered, by excluding a 0.3 nm region from each end of the CNT. E

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Arg+ to ρ̃ ≈ 4 nm−3 or 6 M, as seen in the average values of the green curve. Overall, these results confirm the favorable binding of Arg+ to the CNT surface, and the concomitant enhancement of local density of Arg+, relative to that in the bulk. Potential Mean Force (PMF) of Arg+−CNT, Gdm+− CNT, and Nva−CNT at Infinite Dilution. We used umbrella sampling simulations to quantify water-mediated interactions between the CNT and the various solutes at infinite dilution. A harmonic umbrella potential was placed on the radial coordinate of a selected atom of the solutese.g., the Cα atoms of Arg+ and Nva, or the central carbon atom in the Gdm+ group in Arg+ or in free Gdm+ ions (see Figure 8). At

observed occasional insertion of the aliphatic chain of Nva into one of the ends of the (6,6) CNT, as shown in Figure 6C. Although, in Figures 5A and 5B, there appears to be some curvature dependence of the density profiles (e.g., evident by the differences in numerical values of densities between the outsides of (6,6) and (18,18) CNTs and between the inside and outside of the (18,18) CNT), the overall trend is similar in all cases. As a result, hereafter, we focus on the analysis of densities, interactions, structure, and dynamics primarily near the (6,6) CNT exterior. We also performed the simulations of a (6,6) CNT in solutions of Arg+Cl− sampling a range of concentrations from 0.25 M to ∼2 M. Results for density profiles of Arg+ in these solutions are summarized in Figure 7A. In all solutions, there is

Figure 8. Potential mean force (PMF) between solutes and CNT obtained from umbrella sampling simulations: (A) PMF between Cα carbon and the CNT surface of Arg+ and of Nva, and (B) PMF between the central carbon in the Gdm+ group of free guanidinium and of arginine. The PMFs are set to 0 kJ/mol at rα − rCNT = 1.36 nm in panel (A) and rGdm − rCNT = 1.36 nm in panel (B), for reference.

Figure 7. (A) Equilibrium COM density profile of Arg+ is plotted as a function of r* for increasing molar concentrations of Arg+. (B) Average density (ρ̃0→r* = ∫ r0* ρ(r)rdr/∫ r0* rdr) from 0 nm to r*, where r* = 0.5 or 0.7 nm, as shown by the vertical dashed lines marked “a” and “b”, respectively, in panel (A), is plotted versus the Arg+ concentration.

separation distances greater than ∼1 nm, the PMF is featureless, indicating no significant interactions of Arg+ or Nva with the CNT; however, below 1 nm, a sharp decrease in the Arg+−CNT PMF with a 17 kJ/mol deep minimum at 0.5 nm is observed. In contrast, the interaction of Nva is shorterranged, shows steps, and is significantly less favorable than that for Arg+. Figure 9 makes the molecular details of the favorable interaction of Arg+ with the CNT clear. For separations of >1 nm, Arg+ samples many different orientations freely. However, near 1 nm, the Gdm+ group makes the first contact with the CNT, thus anchoring the Arg+ molecule in a specific orientation. The orientational distribution function obtained at different Arg+−CNT separations also shows this clearly; in the dominant orientations, the Cα−CGdm vector in the Arg+ molecule adopts angles near 10° with the Cα−CNT vector, and the Gdm+ group makes favorable contact with the CNT with its plane parallel to the surface. At shorter separations, other parts of the Arg+ molecule also establish contacts with the CNT, leading to preferred angles of ∼60° (see Figure 9E).

a clearly defined high first peak. The numerical value of the first peak appears to decrease as the bulk concentration of Arg+Cl− increases. At concentrations near and above 1 M, second and third peaks also develop between separations of 0.5 and 1.2 nm, suggesting saturation of the vicinal space and packing of arginine molecules in the second and third shells. Because these density profiles are measured with a bin width of 0.01 nm, they are prone to statistical error. A better measure of the preferential adsorption of solutes can be obtained by integrating the density profile over a larger volume. Figure 7B shows the average densities of Arg+ in cylindrical shells 0.5 and 0.7 nm thick, calculated in this manner. It is clear that the local density of Arg+ in the 0.5-nm-thick cylinder is ρ̃ ≈ 6 nm−3 or 10 M (average values of the red curve) and decreases somewhat as the bulk solution concentration increases. In contrast, in a larger cylinder (0.7 nm thick), there appears to be saturation of F

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Figure 9. Orientational preferences of Arg+ near a (6,6) CNT. Forty (40) equally spaced configurations selected from a 10-ns trajectory are shown. The Arg+ Cα atom is harmonically constrained radially at 1.1, 0.9, and 0.6 nm away from the nanotube surface in panels (A), (B), and (C), respectively. [Note that the scale is different in panels (A), (B), and (C).] CNT is shown in space-fill (cyan), Arg+ is shown in ball-and-stick representation (red, oxygen; white, hydrogen; cyan, carbon; and blue, nitrogen). To highlight the role of the Gdm+ group, it is colored in green. The probability distribution function P(cos θ) of the relative orientation of Arg+, as defined by the angle θ in panel (D), is shown in panel (E).

The importance of the Gdm+ group in mediating interactions with the CNT also becomes clear from Figure 8B, which shows PMFs between the central carbon of the Gdm+ group of Arg+ and of free Gdm+ ion with the CNT surface. Both profiles display a sharp and deep minimum at 0.4 nm. The small additional interactions of non-Gdm+ groups in Arg+ led to the quantitative differences between the two PMFs. Because the PMFs in panels A and B are based on separations of different atoms, one cannot simply add the Nva and Gdm+ PMFs to obtain the Arg+−CNT PMF. Yet, near the contact separation, where both Cα and CGdm+ are approximately the same separation distance from the CNT surface, the summation of the minima values does provide a good estimate of the minimum value of the Arg+−CNT PMF in Figure 8B. Dynamics of Arg+, Gdm+, and Nva at the CNT Surface. We focus on the translational dynamics of Arg+, Gdm+, and Nva molecules in the vicinity of the CNT. The vicinal region is defined by −0.8 ≤ z ≤ 0.8 with z = 0 at the center of the CNT in the axial direction and r − rCNT ≤ 0.5 nm, as shown in the schematic in Figure 10. Furthermore, to account for rapid fluctuations of molecules at the boundary of this region, we used a buffer of 4 ps, to determine if the molecule is in the vicinal region or not. To quantify local translational dynamics for molecules in the vicinal cylindrical shell, we calculate the mean square displacement (MSD) in the rθ direction (see Figure 10B), where the rθ coordinate (appropriately unwrapped in the θ-direction) serves to characterize the essentially one-dimensional (1D) motion of the molecules on the CNT surface. We also performed simulations of dilute aqueous solutions of Arg+, Gdm+, and Nva molecules (10

solutes in 4000 waters) to characterize their diffusion in water. We calculated the self-diffusion coefficient, D, using the Einstein relation, MSD = 2dDt, where d = 1 for motion near the nanotube and d = 3 in bulk water. As shown in Figure 10C, diffusivities of Arg+, Gdm+, and Nva are 1.0, 2.0, and 1.4 ×10−9 m2/s, respectively. These diffusivities are comparable to that of a TIP3P water molecule (5.2 × 10−9 m2/s),72 but are 3−5 times smaller, as expected, based on the larger sizes of these molecules, relative to water. As shown in the previous section, both Arg+ and Gdm+ bind to the CNT rather strongly, with the free energy of binding ranging from 6 kJ/mol to 22 kJ/mol. Despite this strong binding, however, the local translational motion on the CNT surface is not greatly hindered. The local diffusivities for Arg+, Gdm+, and Nva near the CNT surface are 0.3, 1.2, and 1.0 × 10−9 m2/s, respectively. The diffusivity of Arg+ is affected the most and that of Nva is hardly affected, relative to their respective diffusivites in water. The nature of translational dynamics observed here is relatively simple, as a result of the well-defined geometry of the CNT. In more complex environments, such as in the pores of βlactoglobulin crystals or near the surface of a partially hydrated protein in organic media, significant differences were reported between the bulk and local translational dynamics.73,74

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CONCLUDING REMARKS Our molecular dynamics (MD) simulations of lysozyme inside of a single-walled carbon nanotube (CNT) highlight the important role played by arginine ion (Arg+) residues in binding to the surface of a CNT. We investigated this role G

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ASSOCIATED CONTENT

S Supporting Information *

Force field information for arginine (Arg+) and norvaline (Nva), results from simulations of various CNT sizes, and results from simulations of different CNT parameters48 can be found in the Supporting Information. This material is available free of charge via the Internet at http://pubs.acs.org/.

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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS We thank Drs. Hari Acharya, Srivathsan Vembanur, Vasudevan Venkateshwaran, and Siddharth Parimal for helpful discussions and comments on an earlier version of this manuscript. This work was supported by the National Science Foundation (Grant No. CBET-0967937).

Figure 10. (A) Two-dimensional (2D) trajectory of an Arg+ molecule that remains in the vicinity of the CNT over 4 ns (from dark blue (t = 0 ns) to dark red (t = 4 ns)). The vicinal region is highlighted by the red shaded area in the schematic on the left. The rθ (horizontal axis) is obtained by unwrapping the trajectory to remove periodicity in the θ space. The plot thus represents a projection onto a cylindrical plane wrapping the tube. (B) The mean-squared displacement (MSD) on the surface of the CNT based on the unwrapped rθ trajectory shown in panel (A) for the three molecules. (C) The MSD of each molecule in a dilute bulk solution is shown for reference. The self-diffusion coefficient is reported in units of 10−9 m2/s in panels (B) and (C). The error bars in panel (C) are smaller than the marker.

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REFERENCES

(1) Wang, J. Carbon-Nanotube Based Electrochemical Biosensors: A Review. Electroanalysis 2005, 17, 7−14. (2) Karunwi, O.; Baldwin, C.; Griesheimer, G.; Sarupria, S.; Guiseppi-Elie, A. Molecular Dynamics Simulations of Peptide− SWCNT Interactions Related to Enzyme Conjugates for Biosenors and Biofuel Cells. Nano LIFE 2013, 3, 1343007-1−1343007-13. (3) Lewinski, N.; Colvin, V.; Drezek, R. Cytotoxicity of Nanoparticles. Small 2008, 4, 26−49. (4) Karajanagi, S. S.; Vertegel, A. A.; Kane, R. S.; Dordick, J. S. Structure and Function of Enzymes Adsobed onto Single-Walled Carbon Nanotubes. Langmuir 2004, 20, 11594−11599. (5) Wang, S.; Humphreys, E.; Chung, S.; Delduco, D.; Lustig, S.; Wang, H.; Parker, K.; Rizzo, N.; Subramoney, S.; Chiang, Y.; Jagota, A. Peptides with Selective Affinity for Carbon Nanotubes. Nat. Mater. 2003, 2, 196−200. (6) Kam, N. W. S.; Dai, H. Carbon Nanotubes as Intracellular Protein Transporters: Generality and Biological Functionality. J. Am. Chem. Soc. 2005, 127, 6021−6026. (7) Chen, R. J.; Bangsaruntip, S.; Drouvalakis, K. A.; Kam, N. W. S.; Shim, M.; Li, T.; Kin, W.; Utz, P. J.; Dai, H. Noncovalent Functionalization of Carbon Nanotubes for Highly Specific Electronic Biosensors. Proc. Natl. Acad. Sci. U.S.A. 2003, 100, 4984−4989. (8) Ge, C.; Du, J.; Zhao, L.; Wang, L.; Liu, Y.; Li, D.; Yang, Y.; Zhou, R.; Zhao, Y.; Chai, Z.; Chen, C. Binding of Blood Proteins to Carbon Nanotubes Reduces Cytotoxicity. Proc. Natl. Acad. Sci. U.S.A. 2011, 108, 16968−46973. (9) Lin, Y.; Allard, L. F.; Sun, Y.-P. Protein-Affinity of Single-Walled Carbon Nanotubes in Water. J. Phys. Chem. B 2004, 108, 3760−3764. (10) Sang, L.-C.; Coppens, M.-O. Effects of Surface Curvature and Surface Chemistry on the Structure and Activity of Protein Adsorbed in Nanopores. Phys. Chem. Chem. Phys. 2011, 13, 6689−6698. (11) Asuri, P.; Karajanagi, S. S.; Yang, H.; Yim, T.-J.; Kane, R. S.; Dordick, J. S. Increasing Protein Stability Through Control of the Nanoscale Environment. Langmuir 2006, 22, 5833−5836. (12) Asuri, P.; Karajanagi, S. S.; Kane, R. S.; Dordick, J. S. Polymer− Nanotube−Enzyme Composites as Active Antifouling Films. Small 2007, 3, 50−53. (13) Matsuura, K.; Saito, T.; Okazaki, T.; Ohshima, S.; Yumura, M.; Iijima, S. Selectivity of Water-Soluble Proteins in Single-Walled Carbon Nanotube Dispersions. Chem. Phys. Lett. 2006, 429, 497−502. (14) Zorbas, V.; Smith, A. L.; Xie, H.; Ortiz-Acevedo, A.; Dalton, A. B.; Dieckmann, G. R.; Draper, R. K.; Baughman, R. H.; Musselman, I. H. Importance of Aromatic Content for Peptide/Single-Walled

further by simulating solutions of Arg+ and its components, guanidinium (Gdm+) and norvaline (Nva). These simulations show that the strong binding is mediated by the Gdm+ group, which prefers the CNT surface, whereas Nva, which does not contain the Gdm+ group, shows weaker binding. Despite the strong binding of both Arg+ and Gdm+ groups to the CNT, these molecules are relatively dynamic on the CNT surface, displaying a two-dimensional (2D) translational diffusive motion. The relatively strong interactions of Arg+ and Gdm+ groups with CNT open up opportunities for different applications, such as solvating CNTs in water (as demonstrated by Hirano et al.16) or manipulating protein−CNT interactions. For example, site-directed mutagenesis of proteins could be used to add Arg+ residues at specific locations, thus inserting location-specific binding sites. This could allow control over the binding strength to a CNT, as well as control of the protein orientation on a CNT. Such nanobioconjugates may be biologically active and may find applications in composite materials. The competition between residue−CNT interaction versus intrapeptide−protein interactions may be used to control the structure of the peptide/protein on a CNT surface. For a sufficiently strong residue−CNT interaction, alternative interfacial structures may be stabilized on CNT or graphene surfaces. Connecting out results to relevant experiments is also of interest. To that end, atomic force microscopy experiments, such as those by the Walker group75 and the Abbott group76 could be extended to study the interactions of nanotubes or other carbon-based materials with peptides or proteins rich in arginine. H

DOI: 10.1021/la5043553 Langmuir XXXX, XXX, XXX−XXX

Article

Langmuir Carbon Nanotube Interactions. J. Am. Chem. Soc. 2005, 127, 12323− 12328. (15) Calvaresi, M.; Hoefinger, S.; Zerbetto, F. Probing the Structure of Lysozyme−Carbon-Nanotube Hybrids with Molecular Dynamics. Chem.Eur. J. 2012, 18, 4308−4313. (16) Hirano, A.; Tanaka, T.; Kataura, H.; Kameda, T. Arginine Side Chains as a Dispersant for Individual Single-Wall Carbon Nanotubes. Chem.Eur. J. 2014, 20, 4922−4930. (17) He, Z.; Zhou, J. Probing Carbon Nanotube−Amino Acid Interactions in Aqueous Solution with Molecular Dynamics Simulations. Carbon 2014, 78, 500−509. (18) Trinquier, G.; Sanejouand, Y. H. Which Effective Property of Amino Acids Is Best Preserved by the Genetic Code. Protein Eng. 1998, 11, 153−169. (19) Bogan, A. A.; Thorn, K. S. Anatomy of Hot Spots in Protein Interfaces. J. Mol. Biol. 1998, 280, 1−9. (20) Sokalingam, S.; Raghunathan, G.; Soundrarajan, N.; Lee, S.-G. A Study on the Effect of Surface Lysine to Arginine Mutagenesis on Protein Stability and Structure Using Green Fluorescent Protein. PLoS One 2012, 7, No. e40410. (21) Tacke, R.; Chen, Y.; Manley, J. L. Sequence-specific RNA binding by an SR protein requires RS domain phosphorylation: Creation of an SRp40-Specific Splicing Enhancer. Proc. Natl. Acad. Sci. U.S.A. 1997, 94, 1148−1153. (22) Long, J. C.; Caceres, J. F. The SR Protein Family of Splicing Factors: Master Regulators of Gene Expressions. Biochem. J. 2009, 417, 15−27. (23) Sanford, J. R.; Gray, N. K.; Beckmann, K.; Cáceres, J. F. A Novel Role for Shuttling SR Proteins in mRNA Translation. Genes Dev. 2004, 18, 755−768. (24) Tan, R.; Frankel, A. D. Structural Variety of Arginine-Rich RNABinding Peptides. Proc. Natl. Acad. Sci. U.S.A. 1995, 92, 5282−5286. (25) Herce, H. D.; Garcia, A. E.; Litt, J.; Kane, R. S.; Martin, P.; Enrique, N.; Rebolledo, A.; Milesi, V. Arginine-Rich Peptides Destabilize the Plasma Membrane, Consistent with a Pore Formation Translocation Mechanism of Cell-Penetrating Peptides. Biophys. J. 2009, 97, 1917−1925. (26) Huang, K.; Garcia, A. E. Free Energy of Translocating an Arginine-Rich Cell-Penetrating Peptide across a Lipid Bilayer Suggests Pore Formation. Biophys. J. 2013, 104, 412−420. (27) Nepak, D.; Geckeler, K. E. Proteins and Carbon Nanotubes: Close Encounter in Water. Small 2007, 3, 1259−1265. (28) Zheng, M.; Jagota, A.; Semke, E. D.; Diner, B. A.; Mclean, R. S.; Lustig, S. R.; Richardson, R. E.; Tassi, N. G. DNA-Assisted Dispersion and Separation of Carbon Nanotubes. Nat. Mater. 2003, 2, 338−342. (29) Lustig, S. R.; Jagota, A.; Khripin, C.; Zheng, M. Theory of Structure-Based Carbon Nanotube Separations by Ion-Exchange Chromatography of DNA/CNT Hybrids. J. Phys. Chem. B 2005, 109, 2559−2566. (30) Tu, X.; Manohar, S.; Jagota, A.; Zheng, M. DNA sequence motifs for structure-specific recognition and separation of carbon nanotubes. Nature 2009, 460, 250−253. (31) Ao, G.; Khripin, C. Y.; Zheng, M. DNA-Controlled Partition of Carbon Nanotubes in Polymer Aqueous Two-Phase Systems. J. Am. Chem. Soc. 2014, 136, 10383−10392. (32) Liu, J.; Nguyen, M. D. H.; Andya, J. D.; Shire, S. J. Reversible Self-Association Increases the Viscosity of a Concentrated Monoclonal Antibody in Aqueous Solution. J. Pharm. Sci. 2005, 94, 1928−1940. (33) Shukla, D.; Trout, B. L. Interaction of Arginine with Proteins and the Mechanism by Which It Inhibits Aggregation. J. Phys. Chem. B 2010, 114, 13426−13438. (34) Inoue, N.; Takai, E.; Arakawa, T.; Shiraki, K. Arginine and Lysine Reduce the High Viscosity of Serum Albumin Solutions for Pharmaceutical Injection. J. Biosci. Bioeng. 2014, 117, 539−543. (35) Hirano, A.; Kameda, T.; Shinozaki, D.; Arakawa, T.; Shiraki, K. Molecular Dynamics Simulation of the Arginine-Assisted Solubilization of Caffeic Acid: Intervention in the Interaction. J. Phys. Chem. B 2013, 117, 7518−7527.

(36) Hirano, A.; Arakawa, T.; Shiraki, K. Arginine Increases the Solubility of Courmain: Comparison with Salting-In and Salting-Out Additives. J. Biochem. 2008, 144, 363−369. (37) Hirano, A.; Kameda, T.; Arakawa, T.; Shiraki, K. ArginineAssisted Solubilization System for Drug Substances: Solubility Experiment and Simulation. J. Phys. Chem. B 2010, 114, 13455−13462. (38) Ahn, H.; Kim, T.; Choi, H.; Yoon, C.; Um, K.; Nam, J.; Ahn, K. H.; Lee, K. Gelation of Graphene Oxides Induced by Different Types of Amino Acids. Carbon 2014, 71, 229−237. (39) Mason, P. E.; Neilson, G. W.; Enderby, J. E.; Saboungi, M.-L.; Dempsey, C. E.; MacKerell, A. D., Jr.; Brady, J. W. The Structure of Aqueous Guanidinium Chloride Solutions. J. Am. Chem. Soc. 2004, 126, 11462−11470. (40) Das, P. Effect of Cosolvents on Nano-confined Water: A Molecule Dynamics Study. Nanoscale 2012, 4, 2931−2936. (41) Godawat, R.; Jamadagni, S. N.; Garde, S. Unfolding of Hydrophobic Polymers in Guanidinium Chloride Solutions. J. Phys. Chem. B 2010, 114, 2246−2254. (42) Ren, P.; Reichert, D.; He, Q.; Zhang, L.; Tang, H. Understanding the Molecular Dynamics Associated with Polymorphic Transitions of D,L-Norvaline with Solid-State NMR Methods. J. Phys. Chem. B 2011, 115, 2814−2823. (43) Smith, K. M.; Liao, J. C. An Evolutionary Strategy for Isobutanol Production Strain Development in Escherichia Coli. Metab. Eng. 2011, 13, 674−681. (44) Nandi, P.; Sen, G. P. An Antifungal Substance from a Strain of B. Subtilis. Nature 1953, 172, 871−872. (45) Humphrey, W.; Dalke, A.; Schulten, K. VMDVisual Molecular Dynamics. J. Mol. Graphics 1996, 14, 33−38. (46) Hummer, G.; Rasaiah, J. C.; Noworyta, J. P. Water Conduction through the Hydrophobic Channel of a Carbon Nanotube. Nature 2001, 414, 188−190. (47) Rasaiah, J. C.; Garde, S.; Hummer, G. Water in Nonpolar Confinement: From Nanotubes to Proteins and Beyond. Annu. Rev. Phys. Chem. 2008, 59, 713−740. (48) Werder, T.; Walther, J. H.; Jaffe, R. L.; Halicioglu, T.; Koumoutsakos, P. On the Water−Carbon Interaction of Use in Molecule Dynamics Simulations of Graphite and Carbon Nanotubes. J. Phys. Chem. B 2003, 107, 1345−1352. (49) Falk, K.; Sedlmeier, F.; Joly, L.; Netz, R. R.; Bocquet, L. Molecular Origin of Fast Water Transport in Carbon Nanotube Membranes: Superlubricity versus Curvature Dependent Friction. Nano Lett. 2010, 10, 4067−4073. (50) Elliott, J. A.; Sandler, J. K. W.; Windle, A. H.; Young, R. J.; Shaffer, M. S. P. Collapse of Single-Wall Carbon Nanotubes is Diameter Dependent. Phys. Rev. Lett. 2004, 92, 095501-1−095501-4. (51) Tang, T.; Jagota, A.; Hui, C.-Y.; Glassmaker, N. J. Collapse of Single-Walled Carbon Nanotubes. J. Appl. Phys. 2005, 97, 074310-1− 074310-6. (52) Hornak, V.; Abel, R.; Okur, A.; Strockbine, B.; Roitberg, A.; Simmerling, C. Comparison of Multiple Amber Force Fields and Development of Improved Protein Backbone Parameters. Proteins: Struct., Funct., Bioinf. 2006, 65, 712−725. (53) Cornell, W. D.; Cieplak, P.; Bayly, C. I.; Gould, I. R.; Kenneth, M.; Merz, J.; Gerguson, D. M.; Spellmeyer, D. C.; Fox, T.; Caldwell, J. W.; Kollman, P. A. A Second Generation Force Field for the Simulation of Proteins, Nucleic Acids, and Organic Molecules. J. Am. Chem. Soc. 1995, 117, 5179−5197. (54) Jorgensen, W. L.; Tirado-Rives, J. The OPLS Potential Functions for Proteins, Energy Minimizations for Crystals of Cyclic Peptides and Crambin. J. Am. Chem. Soc. 1988, 110, 1657−1666. (55) Pronk, S.; Páll, S.; Schulz, R.; Larsson, P.; Bjelkmar, P.; Apostolov, R.; Shirts, M. R.; Smith, J. C.; Kasson, P. M.; van der Spoel, D.; Hess, B.; Lindahl, E. GROMACS 4.5: A high-throughput and highly parallel open source molecular simulation toolkit. Bioinformatics 2013, 29, 845−854. (56) Nosé, S. A. Molecular Dynamics Method for Simulations in the Canonical Ensemble. Mol. Phys. 1984, 52, 255−268. I

DOI: 10.1021/la5043553 Langmuir XXXX, XXX, XXX−XXX

Article

Langmuir (57) Parrinello, M.; Rahman, A. Polymorphic Transitions in Single Crystals: A New Molecular Dynamics Method. J. Appl. Phys. 1981, 52, 7182−7190. (58) Essmann, U.; Perera, L.; Berkowitz, M. L.; Darden, T.; Lee, H.; Pedersen, L. G. A smooth particle mesh Ewald method. J. Chem. Phys. 1995, 103, 8577−8593. (59) Allen, M. P.; Tildesley, D. J. Computer Simulation of Liquids; Clarendon Press: Oxford, U.K., 1987. (60) Hess, B. P-LINCS: A Parallel Linear Constraint Solver for Molecular Simulation. J. Chem. Theory Comput. 2008, 4, 116−122. (61) Roux, B. The Calculation of the Potential of Mean Force Using Computer Simulations. Comput. Phys. Commun. 1995, 91, 275−282. (62) Zhou, H.-X.; Dill, K. A. Stabilization of Proteins in Confined Spaces. Biochemistry 2001, 40, 11289−11293. (63) Lucent, D.; Vishal, V.; Pande, V. S. Protein Folding Under Confinement: A Role for Solvent. Proc. Natl. Acad. Sci. U.S.A. 2007, 104, 10430−10434. (64) Cheung, M. S.; Thirumalai, D. Effects of Crowding and Confinement on the Structure of the Transition State Ensemble in Proteins. J. Phys. Chem. B 2007, 111, 8250−8257. (65) Mittal, J.; Best, R. B. Thermodynamics and Kinetics of Protein Folding Under Confinement. Proc. Natl. Acad. Sci. U.S.A. 2008, 105, 20233−20238. (66) Tian, J.; Garcia, A. E. Simulation Studies of Protein Folding/ Unfolding Equilibrium under Polar and Nonpolar Confinement. J. Am. Chem. Soc. 2011, 138, 15157−15164. (67) Radhakrishna, M.; Grimaldi, J.; Belfort, G.; Kumar, S. K. Stability of Proteins Inside a Hydrophobic Cavity. Langmuir 2013, 29, 8922−8928. (68) Takaiwa, D.; Hatano, I.; Koga, K.; Tanaka, H. Phase Diagram of Water in Carbon Nanotubes. Proc. Natl. Acad. Sci. U.S.A. 2007, 105, 39−43. (69) Giovambattista, N.; Rossky, P. J.; Debenedetti, P. G. Computational Studies of Pressure, Temperature, and Surface Effect on the Structure and Thermodynamics of Confined Water. Annu. Rev. Phys. Chem. 2012, 63, 179−200. (70) Kubiak-Ossowska, K.; Mulheran, P. A. Multiprotein Interactions during Surface Adsorption: A Molecular Dynamics Study of Lysozyme Aggregation at a Charged Solid Surface. J. Phys. Chem. B 2011, 115, 8891−8900. (71) Kubiak-Ossowska, K.; Mulheran, P. A. Mechanism of Hen Egg White Lysozyme Adsorption on a Charged Solid Surface. Langmuir 2010, 26, 15954−15965. (72) Mark, P.; Nilsson, L. Structure and Dynamics of the TIP3P, SPC, and SPC/E Water Models at 298 K. J. Phys. Chem. A 2001, 105, 9954−9960. (73) Malek, K.; Odijk, T.; Coppens, M.-O. Diffusion of Water and Sodium Counter-ions in Nanopores of a β-lactoglobulin crystal: A molecular dynamics study. Nanotechnology 2005, 16, S522−S530. (74) Yang, L.; Dordick, J. S.; Garde, S. Hydration of Enzyme in Nonaqueous Media is Consistent with Solvent Dependence of Its Activity. Biophys. J. 2004, 87, 812−821. (75) Li, I. T. S.; Walker, G. C. Signature of Hydrophobic Hydration in a Single Polymer. Proc. Natl. Acad. Sci. U.S.A. 2011, 108, 16527− 16532. (76) Ma, C. D.; Wang, C.; Acevedo-Vélez, C.; Gellman, S. H.; Abbott, N. L. Modulation of hydrophobic interactions by proximally immobilized ion. Nature 2015, 517, 347−350.

J

DOI: 10.1021/la5043553 Langmuir XXXX, XXX, XXX−XXX