Assessing Weak Anion Binding to Small Peptides.

Since Hofmeister's seminal studies in the late 19th century, it has been known that salts and buffers can drastically affect the properties of peptides and proteins. These Hofmeister effects can be conceived of in terms of three distinct phenomena/mechanisms: water-salt interactions that indirectly induce the salting-out of a protein by water sequestration by the salt, and direct salt-protein interactions that can either salt-in or salt-out the protein. Unfortunately, direct salt-protein interactions responsible for Hofmeister effects are weak and difficult to quantify. As such, they are frequently construed of as being nonspecific. Nevertheless, there has been considerable effort to better specify these interactions. Here, we use pentapeptides to demonstrate the utility of the H-dimension of nuclear magnetic resonance (NMR) spectroscopy to assess anion binding using N-H signal shifts. We qualify binding using these, demonstrating the upfield shifts induced by anion association and revealing how they are much larger than the corresponding downfield shifts induced by magnetic susceptibility and other ionic strength change effects. We also qualify binding in terms of how the pattern of signal shifts changes with point mutations. In general, we find that the observed upfield shifts are small compared with those induced by anion binding to amide-based hosts, and MD simulations suggest that this is so. Thus, charge-diffuse anions associate mostly with the nonpolar regions of the peptide rather than directly interacting with the amide N-H groups. These findings reveal the utility of 1H NMR spectroscopy for qualifying affinity to peptides─even when affinity constants are very low─and serve as a benchmark for using NMR spectroscopy to study anion binding to more complex systems.

Negative Electron Transfer Collision-Induced Dissociation of G-Quadruplexes: Uncovering the Guanine Radical Anion Loss Pathway.

G-quadruplex (G4) DNA can form highly stable secondary structures in the presence of metal cations, and research has shown its potential as a transcriptional regulator for oncogenes in the human genome. In order to explore the interactions of DNA with metal cations using mass spectrometry, employing complementary fragmentation methods can enhance structural information. This study explores the use of ion-ion reactions for sequential negative electron transfer collision-induced dissociation (nET-CID) as a complement to traditional ion-trap CID (IT-CID). The resulting nET-CID data for G4 anions with and without metal cations show an increase in fragment ion type diversity and yield of structurally informative ions relative to IT-CID. The nET-CID yields greater sequence coverage by virtue of fragmentation at the 3'-side of thymine residues, which is lacking with IT-CID. Potassium adductions to backbone fragments in IT-CID and nET-CID spectra were nearly identical. Of note is a prominent fragment resulting from a loss of a 149 Da anion seen in nET-CID of large, G-rich sequences, proposed to be radical anion guanine loss. Neutral loss of neutral guanine (151 Da) and deprotonated nucleobase loss (150 Da) have been previously reported, but this is the first report of radical anion guanine loss (149 Da). Confirmation of the identity of the 149 Da anion results from the examination of the homonucleobase sequence 5'-GGGGGGGG-3'. Loss of a charged adenine radical anion at much lower relative abundance was also noted for the sequence 5'-AAAAAAAA-3'. DFT modeling indicates that the loss of a nucleobase as a radical anion from odd-electron nucleic acid anions is a thermodynamically favorable fragmentation pathway for G.

Salt-Bridged Peptide Anion Gaseous Structures Enable Efficient Negative Ion Electron Capture Dissociation.

We previously discovered that electron attachment to gaseous peptide anions can occur within a relatively narrow electron energy range. The resulting charge-increased radical ions undergo dissociation analogous to conventional cation electron capture/transfer dissociation (ECD/ETD), thus enabling a novel tandem mass spectrometry (MS/MS) technique that we termed negative ion electron capture dissociation (niECD). We proposed that gaseous zwitterionic structures are required for niECD with electron capture either occurring at or being directed by a positively charged site. Here, we further evaluate this zwitterion mechanism by performing niECD of peptides derivatized to alter their ability to form zwitterionic gaseous structures. Introduction of a fixed positive charge tag, a highly basic guanidino group, or a highly acidic sulfonate group to promote zwitterionic structures in singly charged anions, rescued the niECD ability of a peptide refractory to niECD in its unmodified form. We also performed a systematic study of five sets of synthetic peptides with decreasing zwitterion propensity and found that niECD efficiency decreased accordingly, further supporting the zwitterion mechanism. However, traveling-wave ion mobility-mass spectrometry experiments, performed to gain further insight into the gas-phase structures of peptides showing high niECD efficiency, exhibited an inverse correlation between the orientationally averaged collision cross sections and niECD efficiency. These results indicate that compact salt-bridged structures are also a requirement for effective niECD.

Specific anion effects on urease activity: A Hofmeister study.

The effects of a range of electrolytes on the hydrolysis of urea by the enzyme urease is explored. The autocatalytic behavior of urease in unbuffered solutions and its pH clock reactions are studied. The concentration dependence of the experimental variables is analyzed in terms of specific ion-enzyme interactions and hydration. The results offer insights into the molecular mechanisms of the enzyme, and on the nature of its interactions with the electrolytes. We found that urease can tolerate mild electrolytes in its environment, while it is strongly inhibited by both strong kosmotropic and strong chaotropic anions. This study may cast light on an alternative therapy for Helicobacter pylori infections and contribute to the design of innovative materials and provide new approaches for the modulation of the enzymatic activity.

Computational insight into the effect of alkyl chain length in tetraalkylammonium-based deep eutectic solvents.

The effect of the increase in the alkyl chain length of cation on the properties of deep eutectic solvents based on ethylene glycol has been investigated employing classical molecular dynamics simulations. The change in the structural and dynamic properties in both the bulk and liquid-vapor interface is explored through various analyses. The interaction between the anion and the ethylene glycol increases with an increase in the alkyl chain length of the cation, as observed in the increase of the lifetime of the hydrogen bond formed between the two. The terminal carbon atoms are found to be closer to each other when the cation changes from tetraethylammonium to tetrabutylammonium. The cations are located closer to the interface, and the association of the alkyl chains becomes more significant with increased alkyl chain length, decreasing the surface tension values.

Unveiling the mechanisms behind high CO2 adsorption by the selection of suitable ionic liquids incorporated into a ZIF-8 metal organic framework: A computational approach.

There is growing focus on the crucial task of effectively capturing carbon dioxide (CO2) from the atmosphere to mitigate environmental consequences. Metal-organic frameworks (MOFs) have been used to replace many conventional materials in gas separation, and the incorporation of ionic liquids (ILs) into porous MOFs has shown promise as a new technique for improving CO2 capture and separation. However, the driving force underlying the electronic modulation of MOF nanostructures and the mechanisms behind their high CO2 adsorption remain unclear. This study reports the effect of encapsulating different imidazolium ILs in porous ZIF-8, to clarify the adsorption mechanism of CO2 using density functional theory (DFT)-based approaches. For this purpose, a range of anions, including bis(trifluoromethylsulfonyl)imide [NTf2], methanesulfonate [MeSO3], and acetate [AC], were combined with the 1-ethyl-3-methylimidazolium [EMIM]+ cation. [EMIM]+-based ILs@ZIF-8 composites were computationally investigated to identify suitable materials for CO2 capture. First, the intermolecular and intramolecular interactions between [EMIM]+ and different anions were examined in detail, and their effects on CO2 adsorption were explored. Subsequently, the integration of these ILs into the ZIF-8 solid structure was studied to reveal how their interactions influenced the CO2 adsorption behavior. Our results demonstrate that the incorporation of ILs strongly affects the adsorption capability of CO2, which is highly dependent on the nature of the ILs inside the ZIF-8 framework. DFT simulations further confirmed that the incorporation of ILs into ZIF-8 led to superior CO2 capture compared to isolated ILs and pristine ZIF-8. This improvement was attributed to the mutual interactions between the ILs and ZIF-8, which effectively fine-tuned CO2 adsorption within the composite structure. This understanding may act as a general guide for gaining more insight into the interfacial interactions between ILs and ZIFs structures and how these molecular-level interactions can help predict the selection of ILs for CO2 adsorption and separation, thereby addressing environmental challenges with greater precision and effectiveness.

Universal ion chromatography method for anions in active pharmaceutical ingredients enabled by computer-assisted separation modeling.

Ion Chromatography (IC) is one of the most widely used methods for analyzing ionic species in pharmaceutical samples. A universal IC method that can separate a wide range of different analytes is highly desired as it can save a lot of time for method development and validation processes. Herein we report the development of a universal method for anions in active pharmaceutical ingredients (APIs) using computer-assisted chromatography modeling tools. We have screened three different IC columns (Dionex IonPac AS28-Fast 4 µm, AS19 4 µm and AS11-HC 4 µm) to determine the best suitable column for universal IC method development. A universal IC method was then developed using an AS11-HC 4 µm column to separate 31 most common anionic substances in 36 mins. This method was optimized using LC Simulator and a model which precisely predicts the retention behavior of 31 anions was established. This model demonstrated an excellent match between predicted and experimental analyte retention time (R2 =0.999). To validate this universal IC method, we have studied the stability of sulfite and sulfide analytes in ambient conditions. The method was then validated for a subset of 29 anions using water and organic solvent/water binary solvents as diluents for commercial APIs. This universal IC method provides an efficient and simple way to separate and analyze common anions in APIs. In addition, the method development process combined with LC simulator modeling can be effectively used as a starting point during method development for other ions beyond those investigated in this study.

Polyoxometalates as Potential Artificial Enzymes toward Biological Applications.

Artificial enzymes, as alternatives to natural enzymes, have attracted enormous attention in the fields of catalysis, biosensing, diagnostics, and therapeutics because of their high stability and low cost. Polyoxometalates (POMs), a class of inorganic metal oxides, have recently shown great potential in mimicking enzyme activity due to their well-defined structure, tunable composition, high catalytic efficiency, and easy storage properties. This review focuses on the recent advances in POM-based artificial enzymes. Different types of POMs and their derivatives-based mimetic enzyme functions are covered, as well as the corresponding catalytic mechanisms (where available). An overview of the broad applications of representative POM-based artificial enzymes from biosensing to theragnostic is provided. Insight into the current challenges and the future directions for POMs-based artificial enzymes is discussed.

Selective turn-on sensing of adenosine diphosphate and phosphate anions by ruthenium (II) polypyridine anchored p-tert-butylcalix[4]arene platform.

BACKGROUND: Nucleoside polyphosphate (NPP) anions are important for enzymatic activity and should be monitored by scientists in industry and medicine. By elucidating enzyme kinetics and processes, it aids in the discovery of effective inhibitors and activators. Nucleoside polyphosphate (NPP) anions are used by kinases, GTPases, and glycosyltransferases (GTs). Phosphorylation of certain amino acid residues (Ser, Thr, and Tyr) on proteins requires the breakdown of ATP by protein kinases, which produces ADP. Protein kinases, breakdown of ATP, and NPP are the focus of oncology drug development because the aberrant control of kinase activity is a common cause of cancer. RESULTS: However, a discriminative turn-on fluorescent property is exhibited by non-fluorescent p-tertbutylcalix[4]arene modified 1,2,3-triazole containing bis-ruthenium polypyridyl complex (RL) upon the addition of phosphate anions such as (dihydrogen pyrophosphate (H2P2O72-) and dihydrogen phosphate (H2PO4-)) in CH3CN solvent and Adenosine Diphosphate (ADP) in CH3CN/HEPES (pH = 7.4) buffer (9/1, v/v). The probe RL shows a better-recognizing ability with pyrophosphate anion (H2P2O72-) than dihydrogen phosphate anion (H2PO4-). With H2P2O72- and H2PO4- anions, the RL detection limit was calculated to be as low as 83 nM and 198 nM, respectively. SIGNIFICANCE: The calix[4]arene macrocycle's excellent size and binding cone conformation make it a good host-guest interface for the pyrophosphate anion and ADP. The bis-ruthenium polypyridyl complex's connection to the p-tertbutyl calix[4]arene moiety creates the ADP selectivity turn-on sensor. When moving from mono-nuclear to bi-nuclear ruthenium complex anchored on p-tertbutyl calix[4]arene, the probe can differentiate ADP, ATP, and AMP. Furthermore, this platform is a great resource for creating devices to simultaneously assess phosphate anions in environmental samples.

Quantitatively Differentiating Antibodies Using Charge-State Manipulation, Collisional Activation, and Ion Mobility-Mass Spectrometry.

Antibody-based therapeutics continue to expand both in the number of products and in their use in patients. These heterogeneous proteins challenge traditional drug characterization strategies, but ion mobility (IM) and mass spectrometry (MS) approaches have eased the challenge of higher-order structural characterization. Energy-dependent IM-MS, e.g., collision-induced unfolding (CIU), has been demonstrated to be sensitive to subtle differences in structure. In this study, we combine a charge-reduction method, cation-to-anion proton-transfer reactions (CAPTR), with energy-dependent IM-MS and varied solution conditions to probe their combined effects on the gas-phase structures of IgG1κ and IgG4κ from human myeloma. CAPTR paired with MS-only analysis improves the confidence of charge-state assignments and the resolution of the interfering protein species. Collision cross-section distributions were determined for each of the charge-reduced products. Similarity scoring was used to quantitatively compare distributions determined from matched experiments analyzing samples of the two antibodies. Relative to workflows using energy-dependent IM-MS without charge-state manipulation, combining CAPTR and energy-dependent IM-MS enhanced the differentiation of these antibodies. Combined, these results indicate that CAPTR can benefit many aspects of antibody characterization and differentiation.

Recent progress in the chemistry of heavy aromatics.

The aromaticity and synthetic application of "heavy benzenes", i.e., benzenes containing a heavier Group 14 element (Si, Ge, Sn, and Pb) in place of skeletal carbon, have been the targets of many theoretical and synthetic studies. Although the introduction of a sterically demanding substituent enabled us to synthesize and isolate heavy aromatic species as a stable compound by suppressing their high reactivity and tendency to polymerize, the existence of a protection group is an obstruction to the development of functional materials based on heavy aromatics. This review will delineate the most recent topics in the chemistry of heavy aromatics, i.e., the chemistry of "metallabenzenyl anions", which are the heavier Group 14 element analogs of phenyl anions stabilized by taking advantage of charge repulsion instead of steric protection.

Deciphering the Role of Anions of Ionic Liquids in Modulating the Structure and Stability of ct-DNA in Aqueous Solutions.

Stabilizing biomolecules under ambient conditions can be extremely beneficial for various biological applications. In this context, the utilization of ionic liquids (ILs) in enhancing the stability and preservation of nucleic acids in aqueous solutions is found to be promising. While the role of the cationic moiety of ILs in the said event has been thoroughly explored, the importance of the anionic moiety in ILs, if any, is rather poorly understood. Herein, we examine the function of anions of ILs in nucleic acid stabilization by examining the stability and structure of calf thymus-DNA (ct-DNA) in the presence of various ILs composed of a common 1-ethyl-3-methylimidazolium cations (Emim+) and different anions, which includes Cl-, Br-, NO3-, Ac-,HSO4-and BF4- by employing various spectroscopic techniques as well as Molecular Dynamics (MD) simulation studies. Analysis of our data suggests that the chemical nature of anions including polarity, basicity, and hydrophilicity become an important factor in the overall DNA-IL interaction event. At lower concentrations, the interplay of intermolecular interaction between the IL anions with their respective cations and the solvent molecules becomes a very crucial factor in inducing their stabilizing effect on ct-DNA. However, at higher concentrations of ILs, the ct-DNA stabilization is additionally governed by specific-ion effect. MD simulation studies have also provided valuable insights into molecular-level understanding of the DNA-IL interaction event. Overall, the present study clearly demonstrated that along with the cationic moiety of ILs, the anions of ILs can play a significant role in deciding the stability of duplex DNA in aqueous solution. The findings of this study are expected to enhance our knowledge on understanding of IL-DNA interactions in a better manner and will be helpful in designing optimized IL systems for nucleic acid based applications.

Aromatic pentaamide macrocycles bind anions with high affinity for transport across biomembranes.

The convergent positioning of functional groups in biomacromolecules leads to good binding, catalytic and transport capabilities. Synthetic frameworks capable of convergently locking functional groups with minimized conformational uncertainty-leading to similar properties-are highly desirable but rare. Here we report C5-symmetric aromatic pentaamide macrocycles synthesized in one pot from the corresponding monomers. Their crystal structures reveal a star-shaped, fully constrained backbone that causes ten alternating NH/CH hydrogen-bond donors and five large amide dipoles to orient towards the centre of the macrocycle. With a highly electropositive cavity in a high-energy unbound state, the macrocycles bind anions in a 1:1 stoichiometry in solution, with high affinity for halides and very high affinity for oxoanions. We demonstrate that such macrocycles are able to transport anions across lipid bilayers with a high chloride selectivity and restore the depleted airway surface liquid of cystic fibrosis airway cell cultures.

How a few help all: cooperative crossing of lipid membranes by COSAN anions.

Experimental results show that the presence of a concentration gradient of certain nano-ions (most notably cobaltabisdicarbollide ([o-COSAN]- anions), induce a current across intact artificial phospholipid bilayers in spite of the high Born free energy estimated for these ions. The mechanism underlying this observed translocation of nano-anions across membranes has yet to be determined. Here we show, using molecular dynamics simulations, that the permeation of [o-COSAN]- anions across a lipid bilayer proceeds in a cooperative manner. Single nano-ions can enter the bilayer but permeation is hampered by a free energy barrier of about 8kBT. The interaction between these nano-ions inside a leaflet induces a flip-flop translocation mechanism with the formation of transient, elongated structure inside the membrane. This cooperative flip-flop allows an efficient distribution of [o-COSAN]- anions in both leaflets of the bilayer. These results suggest the existence of a new mechanism for permeation of nano-ions across lipid membranes, relevant for those that have the appropriate self-assembly character.

COSMO-RS guided screening of ionic liquids for the separation of fluorinated greenhouse gases R-410A: Delving into anion, cation effects, and hydrogen bond dynamics.

Reclamation of high-GWP near-azeotropic refrigerant R-410A (50 wt% R-32 (difluoromethane) + 50 wt% R-125 (pentafluoroethane)) can be an effective way to mitigate the greenhouse effect and achieve a circular economy. Efficient ionic liquids (ILs) as extractants needed to be found for the extractive distillation (ED) separation process of R-410A. Given the numerous combinations of cations and anions in ILs, the discovery of an efficient IL via experimental methods proves to be an exceedingly complex task. In this work, the solubilities of R-32, and R-125 in 840 conventional ILs (comprised of 20 cations and 42 anions) were analyzed based on infinite dilution activity coefficient. The absorption mechanisms of R-32 and R-125 in ILs were elucidated by analyzing excess enthalpy (HE), excess Gibbs free energy (GE)) and surface charge density distribution through COSMO-RS (Conductor-like Screening Model for Real Solvents). Results revealed that HE and GE of the mixture formed by R-125 and most ILs surpassed those of R-32, resulting in higher solubility of R-32 in most ILs compared to R-125. Structural changes of anions and cations had a greater effect on the solubility of R-125 in ILs. It is found for the first time that the existence of a strong hydrogen bond donor region in cations/anions generated intense repulsion with the hydrogen atom in R-125. Furthermore, a large area of weak polarity on the surface of cations/anions was difficult to form an effective charge shield with fluorine atoms in R-125, thus inhibiting the dissolution of R-125. Finally based on the identified interaction sites, combined with melting point and viscosity, some novel functional ILs with high selectivity for R-32 + R-125 were designed and determined for actual separation process. These findings significantly enrich the understanding of the solubility mechanism and provide theoretical guidance for designing new ILs for R-410A reclamation.

Anions and Cations Affect Amino Acid Dissociation Equilibria via Distinct Mechanisms.

Salts reduce the pKa of weak acids by a mechanism sensitive to ion identity and concentration via charge screening of the deprotonated state. In this study, we utilize constant pH molecular dynamics simulations to understand the molecular mechanism behind the salt-dependent dissociation of aspartic acid (Asp). We calculate the pKa of Asp in the presence of a monovalent salt and investigate Hofmeister ion effects by systematically varying the ionic radii. We observe that increasing the anion size leads to a monotonic decrease in Asp pKa. Conversely, the cation size affects the pKa nonmonotonically, interpretable in the context of the law of matching water affinity. The net effect of salt on Asp acidity is governed by an interplay of solvation and competing ion interactions. The proposed mechanism is rather general and can be applicable to several problems in Hofmeister ion chemistry, such as pH effects on protein stability and soft matter interfaces.

CO formation from glucose and cellulose as treated in ionic liquids.

The formation of carbon monoxide (CO) from glucose and cellulose by the treatment with various ionic liquids was studied. Ionic liquids with an imidazolium structure as cation and the chloride or acetate as anion were used. Additionally, 1,8-diazabicyclo [5.4.0]undec-7-ene (DBU) was employed as an additive. CO was generated from glucose with a maximum yield of 0.57 mol% after 90 min of treatment at 120 °C in the reaction system in which DBU was added to the ionic liquid. Pyrolysis above 600 °C has been commonly employed for the gasification of lignocellullosics to produce useful gases such as CO. However, this study has revealed that gasification of lignocellullosics to produce CO can occur at significantly lower temperature, specifically at 120 °C.

Solvent Acts as the Referee in a Match-Up Between Charged and Preorganized Receptors.

The prevalence of anion-cation contacts in biomolecular recognition under aqueous conditions suggests that ionic interactions should dominate the binding of anions in solvents across both high and low polarities. Investigations of this idea using titrations in low polarity solvents are impaired by interferences from ion pairing that prevent a clear picture of binding. To address this limitation and test the impact of ion-ion interactions across multiple solvents, we quantified chloride binding to a cationic receptor after accounting for ion pairing. In these studies, we created a chelate receptor using aryl-triazole CH donors and a quinolinium unit that directs its cationic methyl inside the binding pocket. In low-polarity dichloromethane, the 1 : 1 complex (log K1 : 1 ~ 7.3) is more stable than neutral chelates, but fortuitously comparable to a preorganized macrocycle (log K1 : 1 ~ 6.9). Polar acetonitrile and DMSO diminish stabilities of the charged receptor (log K1 : 1 ~ 3.7 and 1.9) but surprisingly 100-fold more than the macrocycle. While both receptors lose stability by dielectric screening of electrostatic stability, the cationic receptor also pays additional costs of organization. Thus even though the charged receptor has stronger binding in apolar solvents, the uncharged receptor has more anion affinity in polar solvents.

AQcalc: A web server that identifies weak molecular interactions in protein structures.

Weak molecular interactions play an important role in protein structure and function. Computational tools that identify weak molecular interactions are, therefore, valuable for the study of proteins. Here, we present AQcalc, a web server (https://aqcalcbiocomputing.com/) that can be used to identify anion-quadrupole (AQ) interactions, which are weak interactions involving aromatic residue (Trp, Tyr, and Phe) ring edges and anions (Asp, Glu, and phosphate ion) both within proteins and at their interfaces (protein-protein, protein-nucleic acids, and protein-lipid bilayer). AQcalc identifies AQ interactions as well as clusters involving AQ, cation-π, and salt bridges, among others. Utilizing AQcalc we analyzed weak interactions in protein models, even in the absence of experimental structures, to understand the contributions of weak interactions to deleterious structural changes, including those associated with oncogenic and germline disease variants. We identified several deleterious variants with disrupted AQ interactions (comparable in frequency to cation-π disruptions). Amyloid fibrils utilize AQ to bury anions at frequencies that far exceed those observed for globular proteins. AQ interactions were detected three and five times more frequently than the hydrogen-bonded AQ (HBAQ) in fibril structures and protein-lipid bilayer interfaces, respectively. By contrast, AQ and HBAQ interactions were detected with similar frequencies in globular proteins. Collectively, these findings suggest AQcalc will be effective in facilitating fine structural analysis. As other web utilities designed to identify protein residue interaction networks do not report AQ interactions, wide use of AQcalc will enrich our understanding of residue interaction networks and facilitate hypothesis testing by identifying and experimentally characterizing these comparably weak but important interactions.

Dynamics in Quaternary Ionic Liquids with Non-Flexible Anions: Insights from Mechanical Spectroscopy.

The present work investigates how mechanical properties and ion dynamics in ionic liquids (ILs) can be affected by ILs' design while considering possible relationships between different mechanical and transport properties. Specifically, we study mechanical properties of quaternary ionic liquids with rigid anions by means of Dynamical Mechanical Analysis (DMA). We are able to relate the DMA results to the rheological and transport properties provided by viscosity, conductivity, and diffusion coefficient measurements. A good agreement is found in the temperature dependence of different variables described by the Vogel-Fulcher-Tammann model. In particular, the mechanical spectra of all the measured liquids showed the occurrence of a relaxation, for which the analysis suggested its attribution to a diffusive process, which becomes evident when the ion dynamics are not affected by the fast structural reorganization of flexible anions on a local level.