Madhusudan Tyagi

Unusual and Highly Tunable Missing-Linker Defects in Zirconium Metal-Organic Framework UiO-66 and Their Important Effects on Gas Adsorption

UiO-66 is a highly important prototypical zirconium metal-organic framework (MOF) compound because of its excellent stabilities not typically found in common porous MOFs. In its perfect crystal structure, each Zr metal center is fully coordinated by 12 organic linkers to form a highly connected framework. Using high-resolution neutron power diffraction technique, we found the first direct structural evidence showing that real UiO-66 material contains significant amount of missing-linker defects, an unusual phenomenon for MOFs. The concentration of the missing-linker defects is surprisingly high, ∼10% in our sample, effectively reducing the framework connection from 12 to ∼11. We show that by varying the concentration of the acetic acid modulator and the synthesis time, the linker vacancies can be tuned systematically, leading to dramatically enhanced porosity. We obtained samples with pore volumes ranging from 0.44 to 1.0 cm(3)/g and Brunauer-Emmett-Teller surface areas ranging from 1000 to 1600 m(2)/g, the largest values of which are ∼150% and ∼60% higher than the theoretical values of defect-free UiO-66 crystal, respectively. The linker vacancies also have profound effects on the gas adsorption behaviors of UiO-66, in particular CO2. Finally, comparing the gas adsorption of hydroxylated and dehydroxylated UiO-66, we found that the former performs systematically better than the latter (particularly for CO2) suggesting the beneficial effect of the -OH groups. This finding is of great importance because hydroxylated UiO-66 is the practically more relevant, non-air-sensitive form of this MOF. The preferred gas adsorption on the metal center was confirmed by neutron diffraction measurements, and the gas binding strength enhancement by the -OH group was further supported by our first-principles calculations.

Dynamics of Biological Macromolecules: Not a Simple Slaving by Hydration Water

We studied the dynamics of hydrated tRNA using neutron and dielectric spectroscopy techniques. A comparison of our results with earlier data reveals that the dynamics of hydrated tRNA is slower and varies more strongly with temperature than the dynamics of hydrated proteins. At the same time, tRNA appears to have faster dynamics than DNA. We demonstrate that a similar difference appears in the dynamics of hydration water for these biomolecules. The results and analysis contradict the traditional view of slaved dynamics, which assumes that the dynamics of biological macromolecules just follows the dynamics of hydration water. Our results demonstrate that the dynamics of biological macromolecules and their hydration water depends strongly on the chemical and three-dimensional structures of the biomolecules. We conclude that the whole concept of slaving dynamics should be reconsidered, and that the mutual influence of biomolecules and their hydration water must be taken into account.

Temperature-dependent dynamical transitions of different classes of amino acid residue in a globular protein.

The temperature dependences of the nanosecond dynamics of different chemical classes of amino acid residue have been analyzed by combining elastic incoherent neutron scattering experiments with molecular dynamics simulations on cytochrome P450cam. At T = 100-160 K, anharmonic motion in hydrophobic and aromatic residues is activated, whereas hydrophilic residue motions are suppressed because of hydrogen-bonding interactions. In contrast, at T = 180-220 K, water-activated jumps of hydrophilic side chains, which are strongly coupled to the relaxation rates of the hydrogen bonds they form with hydration water, become apparent. Thus, with increasing temperature, first the hydrophobic core awakens, followed by the hydrophilic surface.

Diffusion in single supported lipid bilayers studied by quasi-elastic neutron scattering

It seems to be increasingly accepted that the diversity and composition of lipids play an important role in the function of biological membranes. A prime example of this is the case of lipid rafts; regions enriched with certain types of lipids which are speculated to be relevant to the proper functioning of membrane embedded proteins. Although the dynamics of membrane systems have been studied for decades, the microscopic dynamics of lipid molecules, even in simple model systems, is still an active topic of debate. Neutron scattering has proven to be an important tool for accessing the relevant nanometre length scale and nano to picosecond time scales, thus providing complimentary information to macroscopic techniques. Despite their potential relevance for the development of functionalized surfaces and biosensors, the study of single supported membranes using neutron scattering poses the challenge of obtaining relevant dynamic information from a sample with minimal material. Using state of the art neutron instrumentation we were, for the first time, able to model lipid diffusion in single supported lipid bilayers. We find that the diffusion coefficient for the single bilayer system is comparable to the multi-lamellar lipid system. More importantly, the molecular mechanism for lipid motion in the single bilayer was found to be a continuous diffusion, rather than the flow-like ballistic motion reported in the stacked membrane system. We observed an enhanced diffusion at the nearest neighbour distance of the lipid molecules. The enhancement and change of character of the diffusion can most likely be attributed to the effect the supporting substrate has on the lipid organization.

Effects of pressure on the dynamics of an oligomeric protein from deep-sea hyperthermophile

Significance Deep-sea microorganisms can adapt to extreme conditions of high temperature and pressure. What makes these organisms survive and reproduce in such critical conditions remains an open question. Here, we use the quasielastic neutron scattering (QENS) technique to study the dynamic behavior of a hyperthermophilic protein that is found in the deep sea. Our results give evidence that high pressure affects the dynamical properties of proteins by distorting the protein energy landscape in ways that are significantly different for hyperthermophilic and mesophilic proteins. Consequently, a general schematic denaturation phase diagram together with energy landscapes for the two different proteins are derived, and this approach can be used as a general picture to understand the effects of pressure on protein dynamics and activities. Inorganic pyrophosphatase (IPPase) from Thermococcus thioreducens is a large oligomeric protein derived from a hyperthermophilic microorganism that is found near hydrothermal vents deep under the sea, where the pressure is up to 100 MPa (1 kbar). It has attracted great interest in biophysical research because of its high activity under extreme conditions in the seabed. In this study, we use the quasielastic neutron scattering (QENS) technique to investigate the effects of pressure on the conformational flexibility and relaxation dynamics of IPPase over a wide temperature range. The β-relaxation dynamics of proteins was studied in the time ranges from 2 to 25 ps, and from 100 ps to 2 ns, using two spectrometers. Our results indicate that, under a pressure of 100 MPa, close to that of the native environment deep under the sea, IPPase displays much faster relaxation dynamics than a mesophilic model protein, hen egg white lysozyme (HEWL), at all measured temperatures, opposite to what we observed previously under ambient pressure. This contradictory observation provides evidence that the protein energy landscape is distorted by high pressure, which is significantly different for hyperthermophilic (IPPase) and mesophilic (HEWL) proteins. We further derive from our observations a schematic denaturation phase diagram together with energy landscapes for the two very different proteins, which can be used as a general picture to understand the dynamical properties of thermophilic proteins under pressure.

Dynamical and Phase Behavior of a Phospholipid Membrane Altered by an Antimicrobial Peptide at Low Concentration

The mechanism of action of antimicrobial peptides is traditionally attributed to the formation of pores in the lipid cell membranes of pathogens, which requires a substantial peptide to lipid ratio. However, using incoherent neutron scattering, we show that even at a concentration too low for pore formation, an archetypal antimicrobial peptide, melittin, disrupts the regular phase behavior of the microscopic dynamics in a phospholipid membrane, dimyristoylphosphatidylcholine (DMPC). At the same time, another antimicrobial peptide, alamethicin, does not exert a similar effect on the DMPC microscopic dynamics. The melittin-altered lateral motion of DMPC at physiological temperature no longer resembles the fluid-phase behavior characteristic of functional membranes of the living cells. The disruptive effect demonstrated by melittin even at low concentrations reveals a new mechanism of antimicrobial action relevant in more realistic scenarios, when peptide concentration is not as high as would be required for pore formation, which may facilitate treatment with antimicrobial peptides.

Determination of functional collective motions in a protein at atomic resolution using coherent neutron scattering

Coherent neutron scattering determines the forms, time scales, and amplitudes of protein functional collective modes. Protein function often depends on global, collective internal motions. However, the simultaneous quantitative experimental determination of the forms, amplitudes, and time scales of these motions has remained elusive. We demonstrate that a complete description of these large-scale dynamic modes can be obtained using coherent neutron-scattering experiments on perdeuterated samples. With this approach, a microscopic relationship between the structure, dynamics, and function in a protein, cytochrome P450cam, is established. The approach developed here should be of general applicability to protein systems.

Boson Peak in Deeply Cooled Confined Water: A Possible Way to Explore the Existence of the Liquid-to-Liquid Transition in Water

In their Letter, Wang et al. [1] report on an inelastic neutron scattering (INS) experiment where they describe the pressure evolution of a low energy (E ∼ 6 meV) excitation, emerging in confined protonated water only below 230 K at an exchanged momentum Q 1⁄4 2.0 Å−1. Water confinement was used to overcome the unavoidable crystallization occurring below ∼250 K in bulk water. The authors report that a similar finding was also obtained in both bulk (numerical simulations [2]) and confined water at ambient pressure. They refer to this low temperature excitation as a boson peak (BP) [3], and relate its occurrence to the Widom line, concluding that the observed pressure behavior of the BP reveals the signature of the high-density liquid (HDL) to the low-density liquid (LDL) transition proposed [4], though severely questioned [5], for bulk water. We believe these claims to be unconvincing for the following reasons. Comparison with corresponding findings in liquid water.—The authors not only overlook commenting on the actual density of confined liquid water [6–9], but they also neglect to establish any physical relationship with the well known excitations of coherent or incoherent origin occurring at similar energies in bulk liquid water. Since the seminal Raman scattering room temperature studies by Bolla [10], a mode in the ∼5–7 meV range has indeed been regularly observed with optical [11–14], numerical [15], and inelastic x-ray scattering [16–18] and INS techniques [19,20] over a wide thermodynamic range (0–2 kbar, 250–450 K) in both H2O and D2O (see Fig. 4 in Ref. [21]). The microscopic nature of such a mode, underdamped and still well defined at Q 1⁄4 2.0 Å−1, is the subject of controversial single-particle [12,22,23] or collective [21,24] interpretations. Irrespective of its incoherent or coherent nature, this evidence is unquestionable and cannot be ignored. This mode is not easily detectable in high temperature neutron spectra from H2O because of the overwhelming quasielastic contribution. However, its presence always emerges in calculating the hydrogen vibrational density of states, as was done in Ref. [22] at T 1⁄4 256 K, and in bulk or confined H2O from 300 K down to 242 K [25], but not mentioned in Ref. [1]. This mode, but not the BP, was also observed when investigating the vibrational dynamics in amorphous ices [26,27]. Moreover, a bulklike excitation not dependent on temperature was observed down to 205 K in an INS measurement on slightly salty liquid water [28]. Data analysis and treatment.—(i) INS probes at the same time the coherent and incoherent properties of matter with a weight given by their respective neutron cross section and dynamic structure factor. H2O is considered as an incoherent scatterer by reason of the high σinc=σcoh ratio. Yet, this approximation cannot be uncritically adopted as was done in Ref. [1] and a proper estimation of the related ratio SincðQ;ωÞ=ScohðQ;ωÞ at the thermodynamic (P, T) and kinetic (Q;ω) investigated point should be addressed. (ii) An arbitrary interpolating metric is adopted to determine the locus of the BP appearance: the TB parameter is a clumsy, large-error quantity inherent in the slowing down of the thermal diffusion. The peak associated with the low energy—and virtually temperature independent (see Fig. 4 of Ref. [1])—excitation is enhanced by the narrowing of the quasielastic signal upon lowering the temperature. (iii) The exact internal pressure existing in such tiny pores is not directly related to the He applied pressure and is therefore unknown [29]. As a consequence, the confined water phase diagram and properties cannot be unconditionally assigned to those of bulk water. (iv) In order to support the authors’ claims at a less speculative level, the correct BP shape should be determined by calculating the vibrational density of states in excess of that of the corresponding crystalline phase. In conclusion, the whole large body of numeric and experimental investigations on the single particle and collective properties of liquid water report the presence of a weakly dispersing excitation in the 5–7 meV range. We believe that, in order to use the BP as a marker of the HDL or LDL bulk water phases, the authors should perform a more complete data treatment and establish a relation, if any, between the supposed BP peak they observe in a confined environment and the well established bulk mode present in a wide portion of the phase diagram at the same energy.

Water dynamics in rigid ionomer networks

The dynamics of water within ionic polymer networks formed by sulfonated poly(phenylene) (SPP), as revealed by quasi-elastic neutron scattering (QENS), is presented. These polymers are distinguished from other ionic macromolecules by their rigidity and therefore in their network structure. QENS measurements as a function of temperature as the fraction of ionic groups and humidity were varied have shown that the polymer molecules are immobile while absorbed water molecules remain dynamic. The water molecules occupy multiple sites, either bound or loosely constrained, and bounce between the two. With increasing temperature and hydration levels, the system becomes more dynamic. Water molecules remain mobile even at subzero temperatures, illustrating the applicability of the SPP membrane for selective transport over a broad temperature range.

Using the Fluorescence Red Edge Effect to Assess the Long-Term Stability of Lyophilized Protein Formulations

Nanosecond relaxation processes in sugar matrices are causally linked through diffusional processes to protein stability in lyophilized formulations. Long-term protein degradation rates track mean-squared displacement (⟨u(2)⟩) of hydrogen atoms in sugar glasses, a parameter describing dynamics on a time scale of picoseconds to nanoseconds. However, measurements of ⟨u(2)⟩ are usually performed by neutron scattering, which is not conducive to rapid formulation screening in early development. Here, we present a benchtop technique to derive a ⟨u(2)⟩ surrogate based on the fluorescence red edge effect. Glycerol, lyophilized trehalose, and lyophilized sucrose were used as model systems. Samples containing 10(-6) mole fraction of rhodamine 6G, a fluorophore, were excited at either 532 nm (main peak) or 566 nm (red edge), and the ⟨u(2)⟩ surrogate was determined based the corresponding Stokes shifts. Results showed reasonable agreement between ⟨u(2)⟩ from neutron scattering and the surrogate from fluorescence, although deviations were observed at very low temperatures. We discuss the sources of the deviations and suggest technique improvements to ameliorate these. We expect that this method will be a valuable tool to evaluate lyophilized sugar matrices with respect to their ability to protect proteins from diffusion-limited degradation processes during long-term storage. Additionally, the method may have broader applications in amorphous pharmaceutical solids.