Conductometric investigation of nano MnSO4 association parameters and complex formation with glycylglycine in aqueous and methanol-water mixtures: Thermodynamic parameters and potential applications.

This study aimed to investigate the ion pair association values and association parameters of nano MnSO4 in water and methanol-water mixtures (20 % and 40 % methanol by mass percentage) at varying temperatures (298.15, 303.15, 308.15, and 313.15 K) using the conductometric technique. Additionally, the parameters for complex formation between nano MnSO4 and glycylglycine as a ligand were determined. The focus was on elucidating the thermodynamic formation parameters for the nano Mn2+-glycylglycine interaction, with particular emphasis on comparing the 1: 1 and 1: 2 (M: L) complexes to understand the complexation behavior more comprehensively. The results indicated that the complexation process was spontaneous, as evidenced by negative ΔGf (formation free energy change) values, which increased with temperature, highlighting the enhanced spontaneity of the process. The findings provide valuable insights into designing new materials and procedures by enhancing our understanding of the complexation behavior of nano MnSO4 with ligands like glycylglycine, thus contributing to advancements in various applications such as chemical synthesis, medicines, and environmental remediation. By elucidating the thermodynamic aspects of these interactions, the study aimed to provide valuable information that could be utilized in practical applications and further research endeavors.

Prebiotic dimer and trimer peptide formation in gas-phase atmospheric nanoclusters of water.

Insight into the origin of prebiotic molecules is key to our understanding of how living systems evolved into the complex network of biological processes on Earth. By modelling diglycine and triglycine peptide formation in the prebiotic atmosphere, we provide a plausible pathway for peptide growth. By examining different transition states (TSs), we conclude that the formation of diglycine and triglycine in atmospheric nanoclusters of water in the prebiotic atmosphere kinetically favors peptide growth by an N-to-C synthesis of glycines through a trans conformation. Addition of water stabilizes the TS structures and lowers the Gibbs free activation energies. At temperatures that model the prebiotic atmosphere, the free energies of activation with a six water nanocluster as part of the TS are predicted to be 16 kcal mol-1 relative to the prereactive complex. Examination of the trans vs. cis six water transition states reveals that a homodromic water network that maximizes the acceptor/donor nature of the six waters is responsible for enhanced kinetic favorability of the trans N-to-C pathway. Compared to the non-hydrated trans TS, the trans six-water TS accelerates the reaction of diglycine and glycine to form triglycine by 13 orders of magnitude at 217 K. Nature uses the trans N-to-C pathway to synthesize proteins in the ribosome, and we note the similarities in hydrogen bond stabilization between the transition state for peptide synthesis in the ribosome and the transition states formed in nanoclusters of water in the same pathway. These results support the hypothesis that small oligomers formed in the prebiotic atmosphere and rained onto earth's surface.

Formation of Fluorescent Maillard Reaction Intermediates of Peptide and Glucose during Thermal Reaction and Its Mechanism.

The dynamic changes in fluorescence intensity of the Maillard reactions of l-alanyl-l-glutamine (Ala-Gln)/Diglycine (Gly-Gly)/glycyl-l-glutamine (Gly-Gln) and glucose were investigated. It was found that the fluorescence intensity would increase with the reaction time; however, it would decrease after longer heating at higher temperatures, which was accompanied by rapid browning. The strongest intensity occurred at 45, 35, and 35 min at 130 °C for Ala-Gln, Gly-Gly, and Gly-Gln systems, respectively. The simple model reactions of Ala-Gln/Gly-Gly and dicarbonyl compounds were selected to reveal the formation and mechanism of fluorescent Maillard compounds. It was confirmed that both GO and MGO could react with peptides to form fluorescent compounds, especially GO, and this reaction was sensitive to temperature. The mechanism was also verified in the complex Maillard reaction of pea protein enzymatic hydrolysates.

Glycine, Diglycine, and Triglycine Exhibit Different Reactivities in the Formation and Degradation of Amadori Compounds.

A series of Amadori compounds of glucose were prepared from glycine (G-ARP), diglycine (DiG-ARP), and triglycine (TriG-ARP), and identified by UPLC-MS/MS and NMR. The formation rate of ARPs was TriG-ARP > DiG-ARP > G-ARP, and their activation energies were 63.48 kJ/mol (TriG-ARP), 72.84 kJ/mol (DiG-ARP), and 84.76 kJ/mol (G-ARP), respectively, suggesting that ARP was formed more easily from small peptides than from amino acid. Although 1-DG was formed much more difficultly than 3-DG, the same order of the formation of 1-DG, 3-DG, and browning was DiGly > TriGly > Gly. It was also confirmed that more methylglyoxal and glyoxal would be formed from small peptides than equimolar amino acids. Compared with free amino acid, ARP, deoxyglycosones, and their secondary degradation products were more easily formed from dipeptide and tripeptide, thereby stronger browning occurred and higher reactivity was exhibited in Maillard reaction of di- or tripeptide.

Hydration of Simple Model Peptides in Aqueous Osmolyte Solutions.

The biology and chemistry of proteins and peptides are inextricably linked with water as the solvent. The reason for the high stability of some proteins or uncontrolled aggregation of others may be hidden in the properties of their hydration water. In this study, we investigated the effect of stabilizing osmolyte-TMAO (trimethylamine N-oxide) and destabilizing osmolyte-urea on hydration shells of two short peptides, NAGMA (N-acetyl-glycine-methylamide) and diglycine, by means of FTIR spectroscopy and molecular dynamics simulations. We isolated the spectroscopic share of water molecules that are simultaneously under the influence of peptide and osmolyte and determined the structural and energetic properties of these water molecules. Our experimental and computational results revealed that the changes in the structure of water around peptides, caused by the presence of stabilizing or destabilizing osmolyte, are significantly different for both NAGMA and diglycine. The main factor determining the influence of osmolytes on peptides is the structural-energetic similarity of their hydration spheres. We showed that the chosen peptides can serve as models for various fragments of the protein surface: NAGMA for the protein backbone and diglycine for the protein surface with polar side chains.

The Interfacial Interactions of Glycine and Short Glycine Peptides in Model Membrane Systems.

The interactions of amino acids and peptides at model membrane interfaces have considerable implications for biological functions, with the ability to act as chemical messengers, hormones, neurotransmitters, and even as antibiotics and anticancer agents. In this study, glycine and the short glycine peptides diglycine, triglycine, and tetraglycine are studied with regards to their interactions at the model membrane interface of Aerosol-OT (AOT) reverse micelles via 1H NMR spectroscopy, dynamic light scattering (DLS), and Langmuir trough measurements. It was found that with the exception of monomeric glycine, the peptides prefer to associate between the interface and bulk water pool of the reverse micelle. Monomeric glycine, however, resides with the N-terminus in the ordered interstitial water (stern layer) and the C-terminus located in the bulk water pool of the reverse micelle.

Synthesis, Docking, Computational Studies, and Antimicrobial Evaluations of New Dipeptide Derivatives Based on Nicotinoylglycylglycine Hydrazide.

Within a series of dipeptide derivatives (5-11), compound 4 was refluxed with d-glucose, d-xylose, acetylacetone, diethylmalonate, carbon disulfide, ethyl cyanoacetate, and ethyl acetoacetate which yielded 5-11, respectively. The candidates 5-11 were characterized and their biological activities were evaluated where they showed different anti-microbial inhibitory activities based on the type of pathogenic microorganisms. Moreover, to understand modes of binding, molecular docking was used of Nicotinoylglycine derivatives with the active site of the penicillin-binding protein 3 (PBP3) and sterol 14-alpha demethylase's (CYP51), and the results, which were achieved via covalent and non-covalent docking, were harmonized with the biological activity results. Therefore, it was extrapolated that compounds 4, 7, 8, 9, and 10 had good potential to inhibit sterol 14-alpha demethylase and penicillin-binding protein 3; consequently, these compounds are possibly suitable for the development of a novel antibacterial and antifungal therapeutic drug. In addition, in silico properties of absorption, distribution, metabolism, and excretion (ADME) indicated drug likeness with low to very low oral absorption in most compounds, and undefined blood-brain barrier permeability in all compounds. Furthermore, toxicity (TOPKAT) prediction showed probability values for all carcinogenicity models were medium to pretty low for all compounds.

An Isosymmetric High-Pressure Phase Transition in α-Glycylglycine: A Combined Experimental and Theoretical Study.

We investigated the effects of hydrostatic pressure on α-glycylglycine (α-digly) using a combined experimental and theoretical approach. The results of powder X-ray diffraction show a change in compressibility of the axes above 6.7 GPa, but also indicate that the structure remains in the same monoclinic space group, suggesting an isosymmetric phase transition. A noticeable change in the Raman spectra between 6 and 7.5 GPa further supports the observed phase transition. First-principles-based calculations combined with the crystal structure prediction code USPEX predict a number of possible polymorphs at high pressure. An orthorhombic structure with a bent peptide backbone is the lowest enthalpy polymorph above 6.4 GPa; however, it is not consistent with experimental observations. A second monoclinic structure isosymmetric to α-digly, α'-digly, is predicted to become more stable above 11.4 GPa. The partial atomic charges in α'-digly differ from α-digly, and the molecule is bent, possibly indicating different reactivity of α'-digly. The similarity in the lattice parameters predicted from calculations and the axial changes observed experimentally support that the α'-digly phase is likely observed at high pressure. A possible explanation for the isosymmetric phase transition is discussed in terms of relaxing strained hydrogen bonding interactions. Such combined experimental and modeling efforts provide atomic-level insight into how pressure-driven conformational changes alter hydrogen-bonding networks in complicated molecular crystals.

Degronomics: Mapping the Interacting Peptidome of a Ubiquitin Ligase Using an Integrative Mass Spectrometry Strategy.

Human cells make use of hundreds of unique ubiquitin E3 ligases to ensure proteome fidelity and control cellular functions by promoting protein degradation. These processes require exquisite selectivity, but the individual roles of most E3s remain poorly characterized in part due to the challenges associated with identifying, quantifying, and validating substrates for each E3. We report an integrative mass spectrometry (MS) strategy for characterizing protein fragments that interact with KLHDC2, a human E3 that recognizes the extreme C-terminus of substrates. Using a combination of native MS, native top-down MS, MS of destabilized samples, and liquid chromatography MS, we identified and quantified a near complete fraction of the KLHDC2-binding peptidome in E. coli cells. This degronome includes peptides that originate from a variety of proteins. Although all identified protein fragments are terminated by diglycine or glycylalanine, the preceding amino acids are diverse. These results significantly expand our understanding of the sequences that can be recognized by KLHDC2, which provides insight into the potential substrates of this E3 in humans. We anticipate that this integrative MS strategy could be leveraged more broadly to characterize the degronomes of other E3 ligase substrate receptors, including those that adhere to the more common N-end rule for substrate recognition. Therefore, this work advances "degronomics," i.e., identifying, quantifying, and validating functional E3:peptide interactions in order to determine the individual roles of each E3.

Diglycine Enables Rapid Intrabacterial Hydrolysis for Activating Anbiotics against Gram-negative Bacteria.

Antimicrobial drug resistance demands novel approaches for improving the efficacy of antibiotics, especially against Gram-negative bacteria. Herein, we report that conjugating a diglycine (GG) to an antibiotic prodrug drastically accelerates intrabacterial ester-bond hydrolysis required for activating the antibiotic. Specifically, the attachment of GG to chloramphenicol succinate (CLsu) generates CLsuGG, which exhibits about an order of magnitude higher inhibitory efficacy than CLsu against Escherichia coli. Further studies reveal that CLsuGG undergoes rapid hydrolysis, catalyzed by intrabacterial esterases (e.g., BioH and YjfP), to generate chloramphenicol (CL) in E. coli. Importantly, the conjugate exhibits lower cytotoxicity to bone marrow stromal cells than CL. Structural analogues of CLsuGG indicate that the conjugation of GG to an antibiotic prodrug is an effective strategy for accelerating enzymatic prodrug hydrolysis and enhancing the antibacterial efficacy of antibiotics.

UV-vis sensor array combining with chemometric methods for quantitative analysis of binary dipeptide mixture (Gly-Gly/Ala-Gln).

Many endogenous peptides are circulating in bodily fluids at micromole level, and accurate analysis of endogenous peptides at such low level is important. In this study, we presented an extensible, facile and sensitive sensor array based on UV-Vis spectroscopy of the AuNPs combined with chemometric methods for quantitative analysis of binary peptide mixture (Gly-Gly/Ala-Gln) using UV-Vis spectroscopy. High concentration arginine (Arg) and Cr3+ can induce aggregation of the AuNPs and DNA-AuNPs. However, the glycylglycine (Gly-Gly) and alanyl-glutamine (Ala-Gln) can prevent the AuNPs from aggregation. We investigated the prevention of AuNPs aggregation by using Gly-Gly and Ala-Gln mixtures and constructed sensor arrays for quantitative analyses of Gly-Gly and Ala-Gln mixtures. The color change of the solution is relevant to the dose of the target, and it can be visualized by the naked eyes or monitored by UV-Vis spectrometry. Results showed that the concentrations of Arg and Cr3+ are the key factors affecting the sensitivity of the sensor array. Whereas when Gly-Gly and Ala-Gln have to be analyzed simultaneously, concentrations of Arg and Cr3+ both for Gly-Gly and Ala-Gln are difficult to be optimized. Taking the advantages of multivariate analysis and data fusion, PLS models and backward interval PLS (BiPLS) models were built for fused dataset constructed by UV-Vis data obtained at different concentrations of Arg and Cr3+. The best results were obtained from the PLS models. The proposed method can be extended to analysis of other peptides in more complex mixture systems.

Local electronic structure of the peptide bond probed by resonant inelastic soft X-ray scattering.

The local valence orbital structure of solid glycine, diglycine, and triglycine is studied using soft X-ray emission spectroscopy (XES), resonant inelastic soft X-ray scattering (RIXS) maps, and spectra calculations based on density-functional theory. Using a building block approach, the contributions of the different functional groups of the peptides are separated. Cuts through the RIXS maps furthermore allow monitoring selective excitations of the amino and peptide functional units, leading to a modification of the currently established assignment of spectral contributions. The results thus paint a new-and-improved picture of the peptide bond, enhance the understanding of larger molecules with peptide bonds, and simplify the investigation of such molecules in aqueous environment.

Timely Addition of Glutathione for Its Interaction with Deoxypentosone To Inhibit the Aqueous Maillard Reaction and Browning of Glycylglycine-Arabinose System.

The inhibitory effects of glutathione (GSH) and oxiglutathione (GSSG) on Maillard browning were compared, and it was clarified that free sulfhydryl was the key substance for the inhibition. The Amadori rearrangement product (ARP) derived from glycylglycine (Gly-Gly) and arabinose (Ara) was prepared by aqueous Maillard reaction, and LC-MS/MS was used to investigate the reaction products of GSH and purified ARP. Reaction between GSH and deoxypentosone (DP) was found to alter the pathway of aqueous Maillard reaction, which reduced the production of glyoxal, methylglyoxal, and furfural and thereby inhibited the formation of melanoidins. To determine the optimal conditions for browning inhibition, a stepwise increase of temperature was used to prepare Maillard reaction products (MRPs). The results showed that the optimum browning inhibitory effect was obtained by adding GSH after Gly-Gly and Ara heating at 80 °C for 60 min.

Evaluation of New 99mTc(CO)3 + Radiolabeled Glycylglycine In Vivo.

BACKGROUND: Peptide-based agents are used in molecular imaging due to their unique properties, such as rapid clearance from the circulation, high affinity and target selectivity. Many of the radiolabeled peptides have been clinically experienced with diagnostic accuracy. The aim of this study was to investigate in vivo biological behavior of [99mTc(CO)3(H2O)3]+ radiolabeled glycylglycine (GlyGly). METHODS: Glycylglycine was radiolabeled with a high radiolabeling yield of 94.69±2%, and quality control of the radiolabeling process was performed by thin layer radiochromatography (TLRC) and High-Performance Liquid Radiochromatography (HPLRC). Lipophilicity study for radiolabeled complex (99mTc(CO)3-Gly-Gly) was carried out using solvent extraction. The in vivo evaluation was performed by both biodistribution and SPECT imaging. RESULTS: The high radiolabelling yield of 99mTc(CO)3-GlyGly was obtained and verified by TLRC and HPLRC as well. According to the in vivo results, SPECT images and biodistribution data are in good accordance. The excretion route from the body was both hepatobiliary and renal. CONCLUSION: This study shows that 99mTc(CO)3-GlyGly has the potential to be used as a peptide-based imaging agent. Further studies, 99mTc(CO)3-GlyGly can be performed on tumor-bearing animals.

Interfacial rheological behaviors of amphiphilic sodium cholesteryl glycylglycine.

The present study was conducted to investigate the effects of the strong van der Waals interaction and sterol skeleton of surfactants on their interfacial rheological behaviors by comparing the interfacial properties of sodium cholesteryl glycylglycine (Chol-GG-Na) and sodium lauryl glycylglycine (C12-GG-Na) at the oil-aqueous interface. The interfacial dilational rheological experiment results indicate a significant increase in the interfacial activity and intermolecular interaction with the introduction of the cholesteryl group. Therefore, a compact interfacial layer with a remarkably high dilational modulus was obtained with the adsorption of Chol-GG-Na. The cholesteryl group also has a significant impact on the dynamic processes such as it slows down the motion of the molecules due to which the diffusion exchange between the bulk and the interface decreases. Besides, the rigid skeleton makes rearrangement and conformation adjustment difficult. These impacts become more pronounced when the adsorption layer approaches a close and ordered arrangement, which has been confirmed by the relaxation measurements. The reported results provide a theoretical foundation for the potential applications of cholesteryl-based surfactants in the food, pharmaceutical, cosmetic and petroleum industries.

Recognition of the Diglycine C-End Degron by CRL2KLHDC2 Ubiquitin Ligase.

Aberrant proteins can be deleterious to cells and are cleared by the ubiquitin-proteasome system. A group of C-end degrons that are recognized by specific cullin-RING ubiquitin E3 ligases (CRLs) has recently been identified in some of these abnormal polypeptides. Here, we report three crystal structures of a CRL2 substrate receptor, KLHDC2, in complex with the diglycine-ending C-end degrons of two early-terminated selenoproteins and the N-terminal proteolytic fragment of USP1. The E3 recognizes the degron peptides in a similarly coiled conformation and cradles their C-terminal diglycine with a deep surface pocket. By hydrogen bonding with multiple backbone carbonyls of the peptides, KLHDC2 further locks in the otherwise degenerate degrons with a compact interface and unexpected high affinities. Our results reveal the structural mechanism by which KLHDC2 recognizes the simplest C-end degron and suggest a functional necessity of the E3 to tightly maintain the low abundance of its select substrates.

Peptide bond conformation in peptides and proteins probed by dipolar coupling-chemical shift tensor correlation solid-state NMR.

Multidimensional magic-angle spinning solid-state NMR experiments are described that permit cis and trans peptide bonds in uniformly 13C,15N-labeled peptides and proteins to be unambiguously distinguished in residue-specific manner by determining the relative orientations of the amide 13C' CSA and 1H-15N dipolar coupling tensors. The experiments are demonstrated for model peptides glycylglycine and 2,5-diketopiperazine containing trans and cis peptide bonds, respectively. Subsequently, the measurements are extended to two representative proteins that contain exclusively trans peptide bonds, microcrystalline B3 immunoglobulin domain of protein G and Y145Stop human prion protein amyloid fibrils, to illustrate their applicability to a wide range of protein systems.

Electrostatic Effects on the Stability of Peptide Radicals.

We explored the influence of external electric fields (EEFs) on the stability of a glycine dipeptide model radical using high-level quantum chemical methods. Remotely located ions (Cl-/Na+) are used to implement EEF effects. The effects of these ions are reproduced using background point charges and oriented EEFs. Remote charges as far as 900 pm from the Cα radical center can be significantly stabilizing or destabilizing as a function of their relative orientation. The magnitude of these effects is also strongly dependent on the distance between the radical center and the charge location. After examining the strengths and weaknesses of some frequently used quantum mechanics methods in describing these effects properly, a comparison is made on the stability of dipeptide radicals bearing protonable or deprotonable side chains. In this group, the stability of the respective Cα radicals mainly depends on the preferred orientation of the charge-carrying side chain.

Density Functional Studies on Secondary Amides: Role of Steric Factors in Cis/Trans Isomerization.

Cis/trans isomerization of amide bonds is a key step in a wide range of biological and synthetic processes. Occurring through C-N amide bond rotation, it also coincides with the activation of amides in enzymatic hydrolysis. In recently described QM studies of cis/trans isomerization in secondary amides using density functional methods, we highlighted that a peptidic prototype, such as glycylglycine methyl ester, can suitably represent the isomerization and complexities arising out of a larger molecular backbone, and can serve as the primary scaffold for model structures with different substitution patterns in order to assess and compare the steric effect of the substitution patterns. Here, we describe our theoretical assessment of such steric effects using tert-butyl as a representative bulky substitution. We analyze the geometries and relative stabilities of both trans and cis isomers, and effects on the cis/trans isomerization barrier. We also use the additivity principle to calculate absolute steric effects with a gradual increase in bulk. The study establishes that bulky substitutions significantly destabilize cis isomers and also increases the isomerization barrier, thereby synergistically hindering the cis/trans isomerization of secondary amides. These results provide a basis for the rationalization of kinetic and thermodynamic properties of peptides with potential applications in synthetic and medicinal chemistry.

Effective combination therapy of percutaneous ethanol injection and chemotherapy based on injectable low molecular weight gels.

Percutaneous ethanol injection (PEI) therapy was used in liver cancer treatment, however, the therapeutic ethanol in PEI easily flew away from injected solid tumours and hinder the treatment effect. In this paper, injectable supramolecular gels formed by self-assembly of low molecular weight gelators (LMWGs) based on glycylglycine modified phenylboronic acid were prepared to localize ethanol and load chemotherapeutic drug for in situ synergistic therapy. The mechanism, morphology and rheological property of supramolecular gels were characterized by NMR, UV, SEM, etc. The rheological study revealed that the gels were formed in situ rapidly and recovered promptly once damaged. The gels were non-toxicity to both normal 3T3 fibroblasts cells and 4T1 breast cancer cells. Doxorubicin (DOX) hydrochloride and ethanol were encapsulated in the gels for the combination of chemotherapy and PEI therapy. The in vivo anticancer activity of the DOX-loaded gels was carried out in tumour bearing mice. The injected gels were coated around tumour tissues to lock ethanol, and DOX was released sustainingly from the gels to maintain effective concentration to induce the apoptosis of tumour cells. DOX-loaded gels and the ethanol exhibited excellent therapeutic efficacy and low side effects in local cancer therapy.