Combination Of Histidine Research Articles

Antiviral properties of synthetic histidine derivatives containing membranotropic volumetrical carbocycles in their molecule against SARS-CoV-2 virus <i>in vitro</i>

Currently, low molecular-weight compounds are being developed as potential inhibitors of CoVs replication, targeting various stages of the replication cycle, such as major protease inhibitors and nucleoside analogs. Viroporins can be alternative protein targets. The aim of this study is to identify antiviral properties of histidine derivatives with cage substituents in relation to pandemic strain SARS-CoV-2 in vitro. Combination of histidine with aminoadamantane and boron cluster anion [B10H10]2 (compounds IIV) was carried out by classical peptide synthesis. Compound were identified by modern physicochemical methods. Antiviral properties were studied in vitro on a monolayer of Vero E6 cells infected with SARS-CoV-2 (alpha strain) with simultaneous administration of compounds and virus. Derivatives of amino acid histidine with carbocycles and boron cluster were synthesized and their antiviral activity against SARS-CoV-2 was studied in vitro. Histidine derivatives with carbocycles and [B10H10]2 have the ability to suppress virus replication. The solubility of substances in aqueous media can be increased due to formation of hydrochloride or sodium salt. 2HCl*H-His-Rim (I) showed some effect of suppressing replication of SARS-CoV-2 at a viral load of 100 doses and concentration 31.2 g/ml. This is explained by the weakly basic properties of compound I. The presented synthetic compounds showed moderate antiviral activity against SARS-CoV-2. The obtained compounds can be used as model structures for creating new direct-acting drugs against modern strains of coronaviruses.

Facile Synthesis and Characterization of a Bromine-Substituted (Chloromethyl)Pyridine Precursor towards the Immobilization of Biomimetic Metal Ion Chelates on Functionalized Carbons

Multidentate ligands involving tethered pyridyl groups coordinated to transition metal ions have been frequently used to mimic the 3-histidine (3H), 2-histidine-1-carboxylate (2H1C) brace motifs or other combinations of histidine and carboxylate endogenous ligating residues found in bioinorganic metalloenzymes. It is of interest to immobilize these ligand chelates onto heterogeneous supports. This, however, requires the use of bromine-substituted (chloromethyl)pyridines, whose current synthetic routes involve the use of extremely pyrophoric chemicals, such as n-butyllithium that require cryogenic reaction conditions, and toxic chemicals, such as thionyl chloride, that are challenging to handle and require extensive hazard controls. Herein, we report alternative methodologies towards the syntheses of 2-bromo-6-hydroxymethylpyridine and 2-bromo-6-chloromethylpyridine from inexpensive commercially available 2,6-dibromopyridine using isopropylmagnesium chloride lithium chloride complex (Turbo Grignard) and cyanuric chloride which are easier to handle and require milder reaction conditions than the conventional reagents. Gas chromatography-mass spectrometry (GC-MS) methods were developed and simple 1H- and 13C- nuclear magnetic resonance (NMR) and Fourier-transform infrared (FT-IR) spectroscopies were also used to monitor the conversion of both reaction steps and showed that products could be obtained and isolated through simple workups without the presence of unreacted starting material or undesired overchlorinated 2-chloro-6-chloromethylpyridine side product.

Gene Delivery by PAMAM Dendrimer Conjugated with the Nuclear Localization Signal Peptide Derived from Influenza B Virus Nucleoprotein

Polyamidoamine (PAMAM) dendrimer is one of the cationic polymers widely used in nonviral vector-based gene therapy. As PAMAM comprises biodegradable peptide bonds, it shows biocompatibility and relatively low cytotoxicity. In comparison with polyethylenimine (PEI, 25 kDa) that shows high transfection efficiency, the native PAMAM dendrimer has limitations as a gene carrier. To improve the gene expression efficiency, we introduced KRTR peptides derived from the influenza B virus nucleoprotein, known as a nuclear localization signal (NLS), to PAMAM generation 3. We synthesized PAMAM-RTRK and PAMAM-HRTRK and evaluated the effects of the histidine residue on cytotoxicity. The transfection efficiency of PAMAM-RTRK and PAMAM-HRTRK was similar to that of PEI in HepG2 cells, while their gene expression capacities were much higher than capacity of PEI in A549 cells. PAMAM-HRTRK showed relatively lower cytotoxicity than PAMAM-RTRK in the two cell lines, but the transfection capacity was similar for the two polymers. These results imply that the combination of histidine and KRTR peptide may potentially result in a novel PAMAM-based vector that could be used for prolonged gene therapy through the reduction in cytotoxicity and enhancement of gene expression.

Efficient Intracellular Delivery of Cell-Impermeable Cargo Molecules by Peptides Containing Tryptophan and Histidine

We have previously evaluated and reported numerous classes of linear and cyclic peptides containing hydrophobic and hydrophilic segments for intracellular delivery of multiple molecular cargos. Herein, a combination of histidine and tryptophan amino acids were designed and evaluated for their efficiency in intracellular delivery of cell-impermeable phosphopeptides and the anti-HIV drug, emtricitabine. Two new decapeptides, with linear and cyclic natures, both containing alternate tryptophan and histidine residues, were synthesized using Fmoc/tBu solid-phase chemistry. The peptides were characterized and purified by using matrix-assisted laser desorption/ionization (MALDI) spectroscopy and high-performance liquid chromatography (HPLC), respectively. These peptides did not show significant toxicity up to 100 µM in ovarian cancer (SK-OV-3) and leukemia cancer (CCRF-CEM) cells. Furthermore, the cellular uptake of a fluorescence (F’)-labeled cell-impermeable phosphopeptide (F’-GpYEEI) was enhanced in the presence of linear (WH)5 and cyclic [WH]5 by 2- and 8-fold, respectively, compared to the uptake of the phosphopeptide alone. The cellular uptake was not significantly changed in the presence of endocytosis inhibitors. Furthermore, the intracellular uptake of the fluorescently-labeled anti-HIV drug, emtricitabine (F’-FTC), by linear (WH)5 and cyclic [WH]5 in SK-OV-3 cancer cell lines was found to be enhanced by 3.5- and 9-fold, respectively, compared to that of the drug alone. Fluorescent uptake experiments confirmed the localization of F’-GpYEEI-loaded cyclic [WH]5 intracellularly in the SK-OV-3 cancer cell line after 3 h of incubation. Thus, these data demonstrated that [WH]5 containing tryptophan and histidine enhanced the cellular uptake of F’-GpYEEI and emtricitabine.

Combination of histidine, lysine, methionine, and leucine promotes β-casein synthesis via the mechanistic target of rapamycin signaling pathway in bovine mammary epithelial cells.

The ratio of different AA in the diets of cows is vital to improve milk protein yield. β-Casein is one of the important milk proteins with high nutritive value. However, the suitable ratio of essential amino acids (EAA) for the expression of β-casein in the immortalized bovine mammary epithelial cell line is not fully characterized. This study employed response surface methodology to determine the optimal ratio of His, Lys, Met, and Leu on β-casein expression level in vitro and clarified the effect of the 4 EAA on β-casein via the mechanistic target of rapamycin (mTOR) signaling pathway. A central composite design containing 5 axial points per EAA and 28 combinations of the 4 EAA was used in our study. The results of response surface methodology and the changes of the mTOR-related signaling proteins were determined by western blot. The results showed that β-casein level was significantly affected by all 4 EAA (R2 = 0.71). The optimum conditions for β-casein expression are found to be 5.47 mM of His, 7.48 mM of Lys, 1.17 mM of Met, and 8.21 mM of Leu (His:Lys:Met:Leu = 5:6:1:7) in the designed scope of concentration. The interaction of Leu and Met significantly affected β-casein expression (P < 0.01). The phosphorylation of mTOR (Ser2481), regulatory associated protein of target of rapamycin (Ser792), ribosomal protein S6 kinase 1 (Thr389), ribosomal protein S6 (Ser235/236), and eukaryotic elongation factor 2 (Thr56) was increased with the supplementation of either single EAA or an optimal combination of EAA. However, the phosphorylation of eukaryotic initiation factor 4E binding protein 1 (Thr37) was decreased with the addition of Lys, Met, or Leu alone. Furthermore, the phosphorylation (P) of eIF2α (Ser51) was decreased when Met was supplemented alone. Under the optimal mixture of 4 EAA, the phosphorylation of mechanistic target of rapamycin complex 1 signaling proteins was significantly greater than the single EAA supplementations and the expression of β-casein was 98% as high as the positive control (i.e., medium with all AA). A similar trend was found with P-ribosomal protein S6 kinase 1 and P-ribosomal protein S6. In conclusion, the extracellular concentrations of His, Lys, Met, and Leu at a ratio of 5:6:1:7 maximized β-casein expression in the immortalized bovine mammary epithelial cell line may occur via activation of the mechanistic target of rapamycin complex 1 signaling pathway.

0861 Optimal combination of histidine, lysine, methionine, and leucine affect β-casein synthesis via mTOR signaling pathway in bovine mammary epithelial cells

0861 Optimal combination of histidine, lysine, methionine, and leucine affect β-casein synthesis via mTOR signaling pathway in bovine mammary epithelial cells

Secondary interactions involving zinc-bound ligands: Roles in structural stabilization and macromolecular interactions

A large number of proteins contain bound zinc ions. These zinc ions are frequently coordinated by a combination of histidine and cysteine residues. In addition to atoms that coordinate directly to the zinc ions, these side chains have groups that can donate or accept hydrogen bonds from other groups. These secondary interactions can help stabilize the zinc-binding sites, can contribute to protein folding and stability, and, on occasion, can participate in interactions with other macromolecules. Five examples of these secondary interactions are discussed: carbonic anhydrase (where secondary interactions involving histidine residues stabilize the zinc-binding site thermodynamically and kinetically), retroviral nucleocapsid proteins and TRAF proteins (where cysteinate sulfur to peptide NH hydrogen bonds contribute to the structural relationships between adjacent domains), and nucleic acid binding proteins, Zif268 and TIS11 where secondary interactions participate in protein–nucleic acid interactions.

A combinatorial histidine scanning library approach to engineer highly pH‐dependent protein switches

There is growing interest in the development of protein switches, which are proteins whose function, such as binding a target molecule, can be modulated through environmental triggers. Efforts to engineer highly pH sensitive protein-protein interactions typically rely on the rational introduction of ionizable groups in the protein interface. Such experiments are typically time intensive and often sacrifice the protein's affinity at the permissive pH. The underlying thermodynamics of proton-linkage dictate that the presence of multiple ionizable groups, which undergo a pK(a) change on protein binding, are necessary to result in highly pH-dependent binding. To test this hypothesis, a novel combinatorial histidine library was developed where every possible combination of histidine and wild-type residue is sampled throughout the interface of a model anti-RNase A single domain VHH antibody. Antibodies were coselected for high-affinity binding and pH-sensitivity using an in vitro, dual-function selection strategy. The resulting antibodies retained near wild-type affinity yet became highly sensitive to small decreases in pH, drastically decreasing their binding affinity, due to the incorporation of multiple histidine groups. Several trends were observed, such as histidine "hot-spots," which will help enhance the development of pH switch proteins as well as increase our understanding of the role of ionizable residues in protein interfaces. Overall, the combinatorial approach is rapid, general, and robust and should be capable of producing highly pH-sensitive protein affinity reagents for a number of different applications.

99mTc-HYNIC-Gastrin Peptides: Assisted Coordination of 99mTc by Amino Acid Side Chains Results in Improved Performance Both In Vitro and In Vivo

The aim of this study was to determine the effects of assisted coordination by amino acids such as histidine and glutamic acid on the function of (99m)Tc-labeled gastrin peptide-hydrazinonicotinamide (HYNIC) conjugates and their ability to target cholecystokinin-R in small-animal models. Three peptide-HYNIC conjugates containing the -AYGWMDF-NH2 C-terminal sequence and combinations of histidine, glutamic acid, and glycine were synthesized, radiolabeled with (99m)Tc/(99)Tc using either tricine or ethylenediaminediacetic acid as a coligand, and analyzed by the high-performance liquid chromatography and liquid chromatography-mass spectrometric techniques. Stability, receptor binding, and internalization and in vivo targeting in AR42J-bearing mice were assessed. When radiolabeling was performed using tricine as a coligand, the insertion of a histidine residue near the HYNIC residue resulted in the displacement of one molecule of tricine from the coordination sphere, a reduction in the number of radiolabeled species formed, an improvement in the in vitro stability, an increase in the rate of radiopeptide internalization, and a significant improvement in tumor uptake in vivo. When radiolabeling was performed using ethylenediaminediacetic acid as a coligand, no effect on coligand binding, homogeneity, or in vitro stability was observed but a significant improvement in the internalization in vitro and tumor uptake in vivo was again found. All of the complexes formed showed similar receptor affinity in competitive radioligand binding assays. The insertion of histidine into the sequence of peptide-HYNIC conjugates can result in more stable, more homogeneous complexes that show improvements in tumor-targeting performance both in vitro and in vivo.

Dietary influence on quality of farmed Atlantic cod ( Gadus morhua): Effect on glycolysis and buffering capacity in white muscle

The aim was to study the possibility to reduce the pH-drop post mortem in cod muscle by reducing glycolysis or increasing the buffer capacity, hence reducing liquid leakage and fillet gaping during storage. Two feeding experiments were performed with 1-year-old farmed cod (mean weight 0.6 kg–0.9 kg) fed extruded commercial dry feed coated with Experiment 1: sodium oxalate (5 g or 25 g kg − 1), quercetin (5 mg or 25 mg kg − 1), ascorbic acid (1 g or 5 g kg − 1) or magnesium aspartate (5 g or 25 g kg − 1), and Experiment 2: two levels of histidine (12 or 48 g kg − 1), sodium citrate (24 g kg − 1), or a combination of histidine and citrate (12 and 24 g kg − 1, respectively). The cod were kept in duplicate net pens (125 m 3) per dietary treatment. In both experiments, samples were collected for analysis after 1 and 3 weeks of feeding. Dietary supplementation with sodium oxalate, quercetin or magnesium aspartate showed no effect on pH, liquid leakage or gaping, whereas the highest level of ascorbic acid gave an unexpected lower pH and higher liquid leakage during cold storage. These results show that the positive health effects commonly obtained by boosting using high levels of ascorbic acid may be counteracted by reduced fillet quality. The feed containing the highest level of histidine gave a significantly higher content of free histidine, higher muscle pH and lower gaping after 3 weeks of feeding. This indicates that dietary histidine accumulates in cod muscle and increases the buffer capacity post mortem. The liquid leakage from raw muscle was, however, not reduced. Surprisingly, the muscle content of anserine (β-alanyl-π-methyl- l-histidine) and other histidine related compounds (1-methyl-histidine, 3-methyl histidine and carnosine) was not affected by the increased dietary histidine. Dietary supplementation of citrate did not significantly alter post mortem muscle pH, gaping or liquid leakage.

Prolonged rat pancreas preservation using a solution with the combination of histidine and lactobionate.

A newly formulated solution consisting of lactobionate with or without histidine was tested in the preservation of the rat pancreas. Adult male Lewis rats weighing 120-250 g were used as donors and recipients. Fifty-four rat pancreas transplants were performed to investigate the effectiveness of this test solution and to compare it with the standard University of Wisconsin (UW) solution. The final osmolarity of the new test solution was 290-320 mosmol/l. This solution had a higher sodium content and lower potassium content (Na: 110 mEq/l, K: 50 mEq/l). Adenosine, insulin, hydroxyethyl starch and dexamethasone, which are components of the UW solution, were not present in this test solution. Histidine was used as a buffer. Rat pancreases were stored at 4 degrees C in either standard UW solution, or high-Na+-histidine solution, or high-Na+-lactobionate solution for 48 h and 72 h prior to heterotopic transplantation into rats with streptozotocin-induced diabetes mellitus. Functional success rates for rats receiving pancreases that had been preserved in high-Na+-histidine and in high-Na+-lactobionate solutions at 4 degrees C were 100% (5/5) and 100% (7/7) after 48 h preservation, and 50% (4/8) and 14% (1/7) after 72 h preservation, respectively. By contrast, standard UW solution gave only a 44% (4/9) success rate after 48 h preservation and a 0% (0/8) success rate after 72 h preservation. These results demonstrated that the high-Na+-histidine solution was superior to standard UW solution for rat pancreas preservation. This was probably due to the buffer, histidine, which prevented the acidosis of ischemic tissue during the period of preservation.

Prolonged rat pancreas preservation using a solution with the combination of histidine and lactobionate

A newly formulated solution consisting of lactobionate with or without histidine was tested in the preservation of the rat pancreas. Adult male Lewis rats weighing 120–250 g were used as donors and recipients. Fifty-four rat pancreas transplants were performed to investigate the effectiveness of this test solution and to compare it with the standard University of Wisconsin (UW) solution. The final osmolarity of the new test solution was 290–320 mosmol/l. This solution had a higher sodium content and lower potassium content (Na: 110 mEq/l, K: 50 mEq/l). Adenosine, insulin, hydroxyethyl starch and dexamethasone, which are components of the UW solution, were not present in this test solution. Histidine was used as a buffer. Rat pancreases were stored at 4 °C in either standard UW solution, or high-Na +-histidine solution, or high-Na + -lactobionate solution for 48 h and 72 h prior to heterotopic transplantation into rats with streptozotocin-induced diabetes mellitus. Functional success rates for rats receiving pancreases that had been preserved in high-Na*-histidine and in high-Na +-lactobionate solutions at 4°C were 100% (5/5) and 100% (7/7) after 48 h preservation, and 50% (4/8) and 14% (1/7) after 72 h preservation, respectively. By contrast, standard UW solution gave only a 44% (4/9) success rate after 48 h preservation and a 0% (0/8) success rate after 72 h preservation. These results demonstrated that the high-Na +-histidine solution was superior to standard UW solution for rat pancreas preservation. This was probably due to the buffer, histidine, which prevented the acidosis of ischemic tissue during the period of preservation.

EFFECTIVENESS of ORGANIC CHELATORS IN SOLUBILIZING CALCIUM AND ZINC IN FORTIFIED CEREALS UNDER SIMULATED GASTROINTESTINAL pH CONDITIONS

The effectiveness of various chelating agents on the solubilization of zinc and calcium, in a calcium-fortified wheat flake cereal, has been evaluated using an in vitro method. This method is based on solubility during a sequential pH treatment simulating gastrointestinal conditions (endogenous pH of cereal, to pH 2, to pH 7.5). Zinc oxide was added to the cereal at 100% of the U. S. RDA for zinc, alone or in combination with the following organic chelators: citrate, malate, a combination of citrate and malate, lactate and malate, citrate and lactate, glucose, histidine, cysteine, and a combination of histidine and cysteine. the complexation of zinc by citric acid in a 10:1 or 25:1 ligand:zinc molar ratio increased the solubility of both zinc and calcium as compared to the calcium-fortified cereal where zinc was added without ligand. Histidine, cysteine, and the combination of histidine and cysteine enhanced zinc solubility, but did not affect calcium solubility. the other ligands (malate, citrate and malate, lactate and malate, citrate and lactate, glucose) did not have a significant effect on zinc and calcium solubilization.

Resonance Raman scattering from hemoproteins. Effects of ligands upon the Raman spectra of various C-type cytochromes.

Resonance Raman spectra were measured for various C-type cytochromes (mammalian cytochrome c, bacterial cytochrome c3, algal photosynthetic cytochrome f, and alkylated cytochrome c) and a B-type cytochrome (cytochrome b5) in their reduced and oxidized states. (1) For ferrous alkylated cytochrome c, a Raman line sensitive to the replacement of an axial ligand of the heme iron uas found around 1540 cm=1. This ligand-sensitive Raman line indicated the transition from acidic (1545 cm-1) to alkaline (1533 cm-1) forms with pK 7.9. The pH dependence of the Raman spectrum corresponded well to that of the optical absorption spectra. (2) For ferrous cytochrome f, the ligand-sensitive Raman line was found at the same frequency as cytochrome c (1545 cm-1). Accordingly two axial ligands are likely to be histidine and methionine as in cytochrome c. (3) For ferrous cytochrome c3, the frequency of the ligand-sensitive Raman line was between those of cytochrome c and cytochrome b5. Since two axial ligands of the heme iron in cytochrome c3 might be histidines. However, a combination of histidine and methionine as a possible set of two axial ligands was not completely excluded for one or two of the four hemes. (4) In ferrous cytochrome b5, two weak Raman lines appeared at 1302 and 1338 cm-1 instead of the strongest band at 1313 cm-1 of C-type ferrous cytochromes. This suggests the practical use of these bands for the identification of types of cytochromes. The difference in frequency and intensity between B- and C-types of hemes implies that the low effective symmetry of the heme in ferrous cytochrome c is due to vibrational coupling of ring modes with peripheral substituents rather than geometrical disortion of heme.

Structure Versus Metabolic Activities of Amitrole and Its Analogs

Growth of Salmonella typhimurium LT-2 (Loeffler) Cast. and Chalm. was inhibited by 3-amino-l,2,4-triazole (amitrole), and 3-hydroxy-1,2,4-triazole, 3-chloro-l,2,4-triazole, and 5-amino-1,2,3,4-tetrazole (hereinafter referred to respectively as hydroxytriazole, chlorotriazole, and aminotetrazole). Toxicity of 2 × 10−2 M amitrole was partially overcome by 2 × 10−4 M histidine, and more effectively circumvented by combinations of histidine with 2 × 10−4 M adenine, 2 × 10−4 M riboflavin, or 2 × 10−4 M methionine. Serine reduced toxicity of 10−1 M amitrole when combined with both histidine and adenine but was less effective than methionine. Hydroxytriazole inhibition was partially overcome by adenine or less effectively by histidine but not by riboflavin, methionine, or serine. Chlorotriazole and aminotetrazole inhibitions were circumvented by 2 × 10−4 M methionine, 10−3 M serine, and 10−4 M cysteine but not by histidine, adenine, or riboflavin. The toxicity of high concentrations (10−3 M) of cysteine was reduced by chlorotriazole and aminotetrazole.