The Journal of Physical Chemistry B最新文献

Impeded by the complexity of proteinaceous structure and the very weak nature of the noncovalent interactions involved, the detailed mechanisms by which anions induce salting-in Hofmeister effects in proteins and peptides remain unclear. Here, using β-hairpin peptides as models, we examine two approaches to qualify (map) anion binding: ¹H NMR chemical shifts and hydronium-catalyzed hydrogen-deuterium exchange (HDX) rate changes. We demonstrate that each salt investigated─despite an affinity too weak to quantify accurately, caused denaturation to an extent that is both peptide and anion-specific, with more charge-diffuse anions inducing a greater degree of unfolding. Our studies reveal that the HDX mapping provides more detail than chemical shift data. Thus, HDX mapping reveals two slightly different mechanisms of denaturation, depending on the nature of the anion. Namely, assisted by a N-terminal Arg residue, charge-dense Cl^- is chelated by the terminal N-H groups of the hairpin and induces a small degree of denaturation, whereas charge-diffuse anions intercalate deeply into the cation-π-hydrophobic core of the peptide and induce more substantial unfolding. These findings provide a glimpse of the different mechanisms by which anions can induce the salting-in Hofmeister effect in peptides and proteins and suggest HDX as a useful tool to map weak anion binding.

Designing functional molecules which can recognize and modify the activity of a specific protein is a frequently encountered challenge in biology and pharmaceutical chemistry, and requires major effort for each specific protein target. Here we demonstrate that "self-peptides", parts of folded proteins which by their nature are recognizable by the rest of the protein, provide a general route to developing such molecules. Such a synthetic peptide with a chemically prestabilized conformation can incorporate into the target protein during its folding, and can potentially displace its native counterpart to cause functional deficits. This strategy is especially promising for proteins with β-barrel topology, as the seam of the barrel provides a vulnerable target. We demonstrate this strategy by using green fluorescent protein (EGFP) as a model, as its fluorescence is a direct reporter of its conformation and function. Refolding EGFP in the presence of 35 μM of a disulfide-stabilized 20-residue self-peptide (SP1, which resembles a seam, strands 3 and 11, of GFP) quenches the fluorescence by 97%. A peptide with the same composition but a different sequence is only 40% as effective, demonstrating that silencing is relatively specific. Fluorescence correlation spectroscopy and time-resolved fluorescence lifetime measurements show that SP1 causes complete long-term fluorescence silencing of the EGFP molecules it incorporates into. This result can in principle have a biological application if the self-peptide incorporates into a protein during its synthesis, before the nascent protein folds. We show that SP1 can indeed silence nascent sfGFP (closely related to EGFP) during its ribosomal synthesis in an in vitro translation system. Therefore, self-peptides present a potentially general strategy for developing protein-specific silencers for physiological applications.

As important as molecular electrets are for electronic materials and devices, conformational fluctuations strongly impact their macrodipoles and intrinsic properties. Herein, we employ molecular dynamics (MD) simulations with the polarizable charge equilibrium (PQEq) method to investigate the persistence length (L_P) of molecular electrets composed of anthranilamide (Aa) residues. The PQEq-MD dissipates the accepted static notions about Aa macromolecules, and L_P represents the shortest Aa rigid segments. The classical model with a single L_P value does not describe these oligomers. Introducing multiple L_P values for the same macromolecule follows the observed trends and discerns the enhanced rigidity in their middle sections from the reduced stiffness at their terminal regions. Furthermore, L_P distinctly depends on solvent polarity. The Aa oligomers maintain extended conformations in nonpolar solvents with L_P exceeding 4 nm, while in polar media, increased conformational fluctuations reduce L_P to about 2 nm. These characteristics set key guidelines about the utility of Aa conjugates for charge-transfer systems within organic electronics and energy engineering.

In vivo measurement and mapping of oxygen levels within the tissues are crucial in understanding the physiopathological processes of numerous diseases, such as cancer, diabetes, or peripheral vascular diseases. Electron paramagnetic resonance (EPR) associated with biocompatible exogenous spin probes, such as Ox071 triarylmethyl (TAM) radical, is becoming the new gold standard for oxygen mapping in preclinical settings. However, these probes do not show tissue selectivity when injected systemically, and they are not cell permeable, reporting oxygen from the extracellular compartment only. Recently, Ox071-loaded mesoporous silica nanoparticles (MSNs) were proposed for intracellular tumor oxygen mapping in both in vitro and in vivo models. However, the EPR spectrum of the Ox071 spin probe is poorly sensitive to mobility due to the small anisotropy of its g-factor and the absence of hyperfine splitting, making it more difficult to study the mobility of the radical inside the MSNs or its location. Using ¹³C₁ isotopologues of Ox071 and the deuterated Finland trityl (dFT) spin probes, which are highly sensitive to molecular tumbling, we showed that the loading of the probes inside homemade and commercial cationic MSNs drastically decreases their mobility while the high local concentration of the probe inside the MSNs leads to dipolar line width broadening (self-relaxation). This decrease in molecular tumbling and line broadening hampers the oxygen-sensing properties of Ox071 or dFT probes used for EPR oximetry when loaded into MSNs.

Near-interfacial electrons in water can be produced by bombarding an aqueous microjet in vacuum with gas-phase sodium atoms. These Na atoms immediately ionize into Na⁺ and e_s^-, which can then react with surface-active molecules that preferentially populate the surface. We carried out these experiments by reacting e_s^- with the surfactant benzyltrimethylammonium (BTMA⁺) in a 6.7 M LiBr/H₂O microjet at 242 K as a function of pH between 1 and 5. The reaction products, trimethylamine (TMA) and benzyl radical, evaporate into the gas phase where they are detected by a mass spectrometer. We find that TMA evaporation sharply diminishes with increasing H⁺ concentration and is barely visible at pH = 1, while benzyl evaporation varies much less. These results indicate that TMA protonation overwhelms TMA evaporation at 0.1 M H⁺. Diffusion-reaction modeling matches the observed trends and predicts that e_s^- reacts with BTMA⁺ within the top 20 Å at all pH values. However, TMA molecules that evaporate and escape protonation diffuse on average only over 20 Å at pH = 1 but over 1000 Å at pH = 5. These observations emphasize that the near-interfacial region provides a controllable reaction environment that is also an escape route for volatile intermediates, a route that is unavailable deep in the bulk. The competition between evaporation and reaction depends on the solubility of the intermediate, the location of its creation, and the propensity for secondary reactions.

Abnormal amyloid beta (Aβ) aggregation in the form of plaques and its deposition across the human nerve cells are a major hallmark of Alzheimer's disease. Aβ aggregation dynamics and, more importantly, various drugs' effects, either to inhibit the fibril aggregation or to degrade the mature fibrils, have been an area of active research. Large molecule (peptide-based) inhibitors, such as decapeptide (RYYAAFFARR) and pentapeptide (LPFFD), show inhibition/degradation effects on amyloid beta fibrils. Herein, a mathematical model has been proposed. The model simulates Aβ aggregation and inhibitory/degradative action of peptide inhibitors on Aβ fibrillation. Model parameters are tuned by curve fitting the experimental data. The tuned model is used to predict experimental data at different initial dose/fibril concentrations. Model predicted results are observed to be in good agreement with the reported experimental data, demonstrating model's applicability at the molecular level. Sensitivity analyses of the model parameters on the fibril concentration further establish the robustness of the proposed model.

The ERα transcription factor that induces tumor growth is a potential target for breast cancer treatment. Each monomer of the ERα DNA-binding domain (ERαDBD) homodimer has two conserved (Cys)₄-type zinc fingers, ZF1 (N-terminal) and ZF2 (C-terminal). Electrophilic agents release Zn²⁺ by oxidizing the coordinating Cys of the more labile ZF2 to inhibit dimerization and DNA binding. Microsecond-length molecular dynamics (MD) simulations show that greater flexibility of ZF2 in the ERαDBD monomer leaves its Cys more solvent accessible and less shielded from electrophilic attack by sulfur-centered hydrogen bonds than ZF1 which is buried in the protein. In the unreactive DNA-bound dimer, the formation of the dimer interface between the highly flexible D-box motif of ZF2 decreases the solvent accessibility of its Cys toward electrophiles and increases the populations of sulfur-containing hydrogen bonds that reduce their nucleophilicity. Examination of these factors in ERαDBD and other proteins with labile ZF motifs may reveal new targets to treat viral infections and cancer.

Thiocyanate (SCN^–) is known to be a naive ion abundant in biological fluids, blood, and urine. It is also used as a biomarker, as it can penetrate to the brain by crossing the blood brain barrier (BBB) and also gets into the cerebrospinal fluid (CSF) through the blood-CSF barrier. Considering its importance in human physiology, we examine the effect of SCN^– ions on three model proteins: ovalbumin (Ova), bovine serum albumin (BSA), and lysozyme (Lys). We observe that an elevated level of SCN^– (∼0.5 M) leads to an otherwise unusual instant fibrilization of all these proteins at pH 2 at ambient temperature. Field emission scanning electron microscopy (FESEM) and atomic force microscopy (AFM) reveal two distinct initial amyloid-aggregated states: nucleus, protofibril, and two mature fibril states (upon 24 h of incubation): cross-linked network or matrix and bundle-like structures. Despite the structural variation of the three proteins, the formation of these morphologies depends on the counterion: Na⁺ and guanidinium (Gdm⁺). Since these processes are assisted by the associated alteration in protein hydration, we determine individual protein and salt hydration at the thus-obtained different phases using THz-FTIR spectroscopy in the 1.5–22.5 THz (50–750 cm^–1) frequency window. We found that, depending on the counterion, interfacial hydration could act either as a “lubricant” or as a “de-wetting” agent, and the findings can be a potential foundation for future handling of amyloidosis.

Antimicrobial peptides (AMPs) are an alternative source of antibiotics that fight worldwide antibiotic-resistant catastrophes. Dendropsophin 1 (Dc1) is a recently invented novel AMP with 17 amino acid residues obtained from the screen secretion of a frog named Dendropsophus columbianus. Dc1 has two slightly mutated analogues, namely, Dc1.1 and Dc1.2, with improved cationicity and mean amphipathic moment to enhance the selective toxicity against microorganisms. Experimental results indicate that Dc1 and Dc1.1 have similar antimicrobial activity against Gram-negative bacteria Escherichia coli and Gram-positive bacteria Staphylococcus aureus, whereas the synthesized peptide Dc1.2 has shown antimicrobial activity against a wide range of microorganisms. However, the molecular level details of the peptide-membrane interaction and the corresponding changes in the peptide structure remain elusive. In this study, we investigate the bacterial membrane disruption capability of these AMPs by running a total of 14.2 μs long molecular dynamics (MD) simulations. Our findings suggest that all three peptides affect the upper layer of the membrane with different degrees of disruption. After penetration, Dc1 and Dc1.2 retain stable α-helices in the core region, indicating the potential to disrupt the second layer. However, secondary structure analysis shows that Dc1.2 attains extended helical regions on the C-terminus, suggesting it as the superior candidate among the analogues to have the potential of stable pore formation, leading to bacterial cell death. To speed up our study, we adopt a one-transmembrane configuration of Dc1, Dc1.1, and Dc1.2 and find toroidal pores with subsequent water leakage for Dc1.2.

Perceiving a suitably tuned aqueous solution to unravel water's liquid-liquid critical point (LLCP) has become challenging. In this work, we investigated the structures of light and heavy water in the presence of MgCl₂ using excess infrared spectroscopy and density functional theory calculations. The excess spectroscopy enabled us to differentiate the low-density liquid (LDL) water from the other liquid domains of pure water and reveal the new interaction modes between water and the ions. The addition of salt decreases and then increases the population of LDL in aqueous solutions. At the concentrations of 0.4 M in H₂O and 0.6 M in D₂O, the LDL structures undergo the most significant disruption under ambient conditions in the bulk phase. Furthermore, threshold concentrations of 1 and 1.3 M for light and heavy water, respectively, were found to induce higher LDL populations. The current investigation sheds light on the intriguing liquid-liquid phase transition (LLPT) and the LLCP of water.