Phenylalanine Moiety Research Articles

Enzymatically Biodegradable Polyrotaxane-Deferoxamine Conjugates for Iron Chelation.

Chelation therapy is frequently used to help reduce excess iron in the body, but current chelators such as deferoxamine (DFO) are plagued by short blood circulation times, which necessitates infusions and can cause undesirable toxic side effects in patients. To address these issues, polyrotaxanes (PR) were synthesized by threading α-cyclodextrin (α-CD) onto poly(ethylene glycol) bis(amine) (PEG-BA, MW 3400 g/mol) capped with enzymatically cleavable bulky Z-L phenylalanine (Z-L Phe) moieties. The resulting PR was conjugated to DFO and hydroxypropylated to generate the final polyrotaxane-DFO (hPR-DFO). The iron chelating capability of hPR-DFO was verified by UV-vis absorption spectroscopy and the ability of materials to degrade into smaller CD-conjugated DFO fragments (hCD-DFO) in the presence of the protease was confirmed via gel permeation chromatography. In vitro studies in iron-overloaded macrophages reveal that hPR-DFO can significantly reduce the cytotoxicity of the drug while maintaining its chelation efficacy, and that it is more rapidly endocytosed and trafficked to lysosomes of iron-overloaded cells in comparison to non-iron-overloaded macrophages. In vivo studies indicate that iron-overloaded mice treated with hPR-DFO displayed lower serum ferritin levels (a measure of iron burden in the body) and could eliminate excess iron by both the renal and fecal routes. Moreover, there was no gross evidence of acute toxicological damage to the liver or spleen.

Amino-Acid-Incorporating Nonionic Surfactants for Stabilization of Protein Pharmaceuticals.

With the rapid development of protein-based pharmaceutical products over the past decade, one of the biggest challenges in product development is maintaining the structural stability of proteins during purification, processing, and storage. In this work, the design of a new class of surfactants, polyether-modified N-acyl amino acids, is presented. One surfactant from this series, containing a phenylalanine moiety, demonstrated remarkable stabilization against aggregation of several model protein drugs. Dynamic light scattering, size exclusion chromatography, and circular dichroism all show the rate of thermally accelerated protein aggregation slowed. IgG aggregation was reduced by 3-fold compared to polysorbate controls. Testing of Orencia, a prescription biologic drug for rheumatoid arthritis, demonstrated a 36% improvement in monomer retention upon heat-aging.

The Determination of Protonation Constants of Peptidomimetic Cyclophanes in Binary Methanol-Water Mixtures.

The protonation constants of new group of peptidomimetic cyclophanes with valine or phenylalanine moieties incorporated into the macrocyclic skeleton as well as their linear analogues were determined by potentiometric measurements in solutions of methanol-water mixtures at 25°C and constant ionic strength. The influence of cavity size, location of protonation sites, and attached substituents of the macrocyclic ligands on the protonation constants were discussed on the basis of potentiometric measurement as well as H1-NMR results.

Synthesis and carbonic anhydrase inhibitory properties of amino acid – coumarin/quinolinone conjugates incorporating glycine, alanine and phenylalanine moieties

N-Protected amino acids (Gly, Ala and Phe) were reacted with amino substituted coumarin and quinolinone derivatives, leading to the corresponding N-protected amino acid–coumarin/quinolinone conjugates. The carbonic anhydrase (CA, EC 4.2.1.1) inhibitory activity of the new compounds was assessed against various human (h) isoforms, such as hCA I, hCA II, hCA IV and hCA XII. The quinolinone conjugates were inactive as enzyme inhibitors, whereas the coumarins were ineffective hCA I/II inhibitors (KIs > 50 μM) but were submicromolar hCA IV and XII inhibitors, with inhibition constants ranging between 92 nM and 1.19 μM for hCA IV, and between 0.11 and 0.79 μM for hCA XII. These coumarin derivatives, as many others reported earlier, thus show an interesting selective inhibitory profile for the membrane-bound over the cytosolic CA isoforms.

Biomimetic triblock and multiblock copolymers containing l‐Phenylalanine moieties showing healing and enhanced mechanical properties

ABSTRACTTriblock and multiblock copolymers of methyl acrylate containing short blocks of the hydrogen bonding monomer N‐acryloyl‐l‐phenylalanine were prepared via reversible addition–fragmentation chain transfer polymerization in two steps using a bifunctional trithiocarbonate for the triblock copolymer and a polyfunctional trithiocarbonate for the multiblock copolymer. The polymer materials were investigated via tensile testing showing that the hydrogen bonding monomer induces a pronounced increase in toughness. The toughness of the material is further enhanced when going from triblock to multiblock topology. Both types of copolymer display a very strong healing effect, with the samples' toughness (which is increased by drawing) becoming even larger after breaking and healing. Already, a very small content of only 0.1 mol % of N‐acryloyl‐l‐phenylalanine improves the mechanical properties of these thermoplastic elastomers significantly. © 2015 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2015, 53, 2809–2819

Direct amidation of the phenylalanine moiety in short peptides via Pd-catalyzed C–H activation/C–N formation

The direct amidation of dipeptides through direct aryl C–H transformation of the phenylalanine residue via Pd catalysis.

Prolific cluster emission in sputtering of phenylalanine by argon-cluster ion bombardment

Large Arn+ cluster ions (with n∼1500 Ar atoms per cluster) with a bombarding energy of 10keV were used to investigate the sputter-induced emission of positive secondary ions from a phenylalanine specimen by orthogonal time-of-flight SIMS. An abundant flux of phenylalanine cluster ions (Mn+H)+ with n≤12 was observed. The yield of dimers relative to monomers is found to amount to 50 − 60% whereas that of trimers and tetramers is roughly 10%. Tentatively, this prolific formation of these cluster species can then be ascribed to the concerted action of the large number of Ar atoms within their impact zone at the surface: these low-energy Ar species (with an average energy of only few eV) may effect the soft cleavage of the phenylalanine bonds in the solid and lead, eventually, to the intact emission of these phenylalanine moieties.

Effect of the 4′-substituted phenylalanine moiety of sansalvamide A peptide on antitumor activity

Eight sansalvamide A peptide analogues with 4′-fluoride, 4′-chloride, 4′-bromide, 4′-iodide, and 4′-methoxyphenylalanine moieties were synthesized. The effect of these para-substitutions of sansalvamide A peptide on their cytotoxicity was evaluated using HCT-116, MDA-MB-231, HT-29, HCT-15, K562, HeLa, and A549 cell lines. The 4′-methoxyphenylalanine analog of sansalvamide A peptide was found to be a promising antitumor agent.

2′-Fluoro-6′-methylene-carbocyclic adenosine phosphoramidate (FMCAP) prodrug: In vitro anti-HBV activity against the lamivudine–entecavir resistant triple mutant and its mechanism of action

Novel 2′-fluoro-6′-methylene-carbocyclic adenosine (FMCA) monophosphate prodrug (FMCAP) was synthesized and evaluated for its in vitro anti-HBV potency against a lamivudine–entecavir resistant clone (L180M+M204V+S202G). FMCA demonstrated significant antiviral activity against wild-type as well as lamivudine–entecavir resistant triple mutant (L180M+M204V+S202G). The monophosphate prodrug (FMCAP) demonstrated greater than 12-fold (12×) increase in anti-HBV activity without increased cellular toxicity. Mitochondrial and cellular toxicity studies of FMCA indicated that there is no significant toxicity up to 100μM. Mode of action studies by molecular modeling indicate that the 2′-fluoro moiety by hydrogen bond as well as the Van der Waals interaction of the carbocyclic ring with the phenylalanine moiety of the polymerase promote the positive binding, even in the drug-resistant mutants.

Prohormone-substrate peptide sequence recognition by peptidylglycine α-amidating monooxygenase and its reflection in increased glycolate inhibitor potency

The interactions of nineteen peptide substrates and fifteen analogous peptidomimetic glycolate inhibitors with human peptidylglycine α-amidating monooxygenase (PAM) have been investigated. The substrates and inhibitors are the prohormones of calcitonin and oxytocin and their analogues. PAM both secreted into the medium by and extracted from DMS53 small lung carcinoma cells has been studied. The results show that recognition of the prooxytocin and procalcitonin peptide sequences by the enzyme extends more than four and five amino acid residues, respectively, from their C-termini. This substrate sequence recognition is mirrored by increased inhibitor potency with increased peptide length in the glycolate peptidomimetics. Substitution of the C-terminal penultimate glycine and proline residues of prooxytocin and procalcitonin and their analogues with phenylalanine increases the enzyme binding affinity. However, this changes the binding mode from one that depends on peptide sequence recognition to another primarily determined by the phenylalanine moiety, for both the substrates and analogous glycolate inhibitors.

Use of silane coupling agent for surface modification of zinc oxide as inorganic filler and preparation of poly(amide-imide)/zinc oxide nanocomposite containing phenylalanine moieties

A series of novel poly(amide–imide)/ZnO nanocomposites with modified ZnO nanoparticles contents was prepared by ultrasonic irradiation. For this purpose, surface of ZnO nanoparticle was modified with $\boldsymbol\gamma$ -aminopropyltriethoxysilane as a coupling agent. Then the effect of surface modification on dispersion of nanoparticles, thermal stability and UV absorption property of the obtained nanocomposites were investigated. The resulting novel nanocomposites were characterized by several techniques. Field emission scanning electron microscopy and transmission electron microscopy analyses of the nanocomposites were performed in order to study the dispersion of nanofillers in the polymer matrix. According to thermogravimetry analysis results, the addition of ZnO nanoparticles improved thermal stability of the obtained nanocomposites. Since the resulting nanocomposites contain phenylalanine amino acid and ZnO, they are expected to be biocompatible as well as biodegradable.

Discovery and Synthesis of Namalide Reveals a New Anabaenopeptin Scaffold and Peptidase Inhibitor

The discovery, structure elucidation, and solid-phase synthesis of namalide, a marine natural product, are described. Namalide is a cyclic tetrapeptide; its macrocycle is formed by only three amino acids, with an exocyclic ureido phenylalanine moiety at its C-terminus. The absolute configuration of namalide was established, and analogs were generated through Fmoc-based solid phase peptide synthesis. We found that only natural namalide and not its analogs containing l-Lys or l-allo-Ile inhibited carboxypeptidase A at submicromolar concentrations. In parallel, an inverse virtual screening approach aimed at identifying protein targets of namalide selected carboxypeptidase A as the third highest scoring hit. Namalide represents a new anabaenopeptin-type scaffold, and its protease inhibitory activity demonstrates that the 13-membered macrolactam can exhibit similar activity as the more common hexapeptides.

Discovery of a sensitive Cu(ii)-cyanide “off–on” sensor based on new C-glycosyl triazolyl bis-amino acid scaffold

A new functional glycosyl peptidomimetic, featuring a C-glucosyl 1,4-dimethoxynaphthalene backbone in conjugation with two triazolyl phenylalanine moieties on its adjacent C3,4-positions, was readily synthesized via click chemistry. Primary optical measurements indicated that the fluorescence of the ester form of this probe (4) could be selectively quenched by Pb(2+). In contrast, the fluorescence intensity of its analog 5 with released carboxylic groups was uniquely diminished by Cu(2+) with remarkably enhanced sensitivity and selectivity. Moreover, subsequent addition of cyanide to the methanol solution of the resulting Cu(2+)-5 complex induced its fluorescence recovery with a nanomolar detection limit, which was two orders of magnitude smaller than the regulated concentration limit of CN(-) in drinking water. This suggests the promising applicability of C-glycosyl bis-triazolyl amino acid scaffold in the future design and exploration of sensitive "off-on" Cu(II)-cyanide chemosensors.

Strongly reduced fragmentation and soft emission processes in sputtered ion formation from amino acid films under large Arn+ (n ≤ 2200) cluster ion bombardment

The analysis of organic and biological substances by secondary-ion mass spectrometry (SIMS) has greatly benefited from the use of cluster ions as primary bombarding species. Thereby, depth profiling and three-dimensional (3D) imaging of such systems became feasible. Large Ar(n)(+) cluster ions may constitute a further improvement in this direction. To explore this option, size-selected Ar(n)(+) cluster ions with 300 ≤ n ≤ 2200 (bombarding energies 5.5 and 11 keV) were used to investigate the emission of positive secondary ions from four amino acid specimens (arginine, glycine, phenylalanine, and tyrosine) by time-of-flight SIMS. For all cluster sizes, the protonated molecule of the respective amino acid is observed in the mass spectra. With increasing cluster size the number of fragment ions decreases strongly in relation to the intact molecules, to the extent that the fraction of fragment ions amounts to less than 10% in some cases. Such 'soft' emission processes also lead the ejection of dimers and even multimers of the amino acid molecules. In the case of the phenylalanine, secondary ion species composed of up to at least seven phenylalanine moieties were observed. Tentatively, the ionization probability of the emitted molecules is envisaged to depend on the presence of free protons in the emission zone. Their number can be expected to decrease concurrently with the decreasing amount of fragmentation for large Ar(n)(+) cluster ions (i.e. for low energies per cluster atom).

Antiviral activity of novel 2′-fluoro-6′-methylene-carbocyclic adenosine against wild-type and drug-resistant hepatitis B virus mutants

Novel 2'-fluoro-6'-methylene-carbocyclic adenosine (9) was synthesized and evaluated its anti-HBV activity. The titled compound demonstrated significant antiviral activity against wild-type as well as lamivudine, adefovir and double lamivudine/entecavir resistant mutants. Molecular modeling study indicate that the 2'-fluoro moiety by a hydrogen bond, as well as the van der Waals interaction of the carbocyclic ring with the phenylalanine moiety of the polymerase promote the positive binding, even in the drug resistant mutants.

Synthesis of 4-(2-hydroxyaryl)-3-nitro-4 H-chromenes

A combinatorial library of 4-(2-hydroxyaryl)-3-nitro-4 H-chromenes was synthesized in high yield by C4-SMe substitution in N-alkyl/phenyl 4-(methylthio)-3-nitro-4 H-chromen-2-amines with a variety of phenols. The reaction always provided C2 substitution in the phenol ring, dictated by hydrogen bond interactions between the phenolic hydroxyl group and the nitro group in 3-nitro-4 H-chromenes. Reduction of the nitro group with concomitant hydrolysis of the enamine in 4-(2-hydroxyaryl)-3-nitro-4 H-chromenes with Zn, Ac 2O in AcOH furnished hybrid amino-acid lactone incorporating ortho-tyrosine and phenyl alanine moieties.

Pluripotentialities of a quenched fluorescent peptide substrate library: enzymatic detection, characterization, and isoenzymes differentiation

Protease inhibitors represent a major class of drugs, even though a large number of proteases remain unexplored. Consequently, a great interest lies in the identification of highly sensitive substrates useful for both the characterization and the validation of these enzyme targets and for the design of inhibitors as potential therapeutic agents through high-throughput screening (HTS). With this aim, a synthetic substrate library, in which the highly fluorescent ( L)-pyrenylalanine residue (Pya) is efficiently quenched by its proximity with the p-nitro-( L)-phenylalanine (Nop) moiety, was designed. The cleavage between Pya and Nop leads to a highly fluorescent metabolite providing the required sensitivity. This library, characterized by a water-soluble primary sequence Ac-SGK-Pya-(X) n -Nop-GGK-NH 2, X being a mixture of 10 natural amino acids (A, I, L, K, F, W, E, Q, T, P) and n varying from 0 to 3, was validated using enzymes belonging to the four main types of hydrolases: serine-, metallo-, cystein-, and aspartyl-proteases. The selectivity of substrates belonging to this library was evidenced by characterizing specific substrates for the isoenzymes NEP-1 and NEP-2. This library easily synthesized is of great interest for the identification and development of selective and specific substrates for still uncharacterized endoproteases.

Chiral recognition of macromolecules with cyclodextrins: pH- and thermosensitive copolymers from N-isopropylacrylamide and N-acryloyl-D/L-phenylalanine and their inclusion complexes with cyclodextrins

In the present work we report the enantioselective recognition of water soluble stimuli-responsive polymers bearing phenylalanine moieties via host-guest interaction with β-cyclodextrin and randomly-methylated-β-cyclodextrin (RAMEB-CD). We synthesised N-acryloyl-D/L-phenylalanine monomers (2D, 2L) which were then copolymerised under free radical conditions with N-isopropylacrylamide (NIPAAm). The resulting copolymers 3D and 3L exhibit a lower critical solution temperature (LCST) of 25 °C. As a further benefit, the presence of a free carboxylic group in the copolymer system gives a high sensitivity to the pH value in respect to the LCST value. The enantioselective recognition of the side groups of copolymers 3D and 3L and their solubility behaviour were investigated by dynamic light scattering and 2D NMR spectroscopy, respectively.

Aromatic Amino Acid Monolayers Sandwiched between Gold and Silver: A Combined Tip-Enhanced Raman and Theoretical Approach

Tip-enhanced Raman scattering (TERS) measurements and theoretical calculations of the aromatic amino acids phenylalanine, tyrosine, and tryptophan sandwiched between gold and silver layers are presented and discussed. Atomically flat micrometer-sized gold nanoplates were chosen as substrates and covered with a monolayer of the respective amino acid. On the basis of spectral data, the carboxylate and amino groups were determined to be the preferential moieties attached to the metals in the cavity. The experimental observation of benzene rings oriented parallel to the gold surface and the influence of the silver tip are discussed and compared with theoretical calculations. Finally, it is shown that TERS allows a clear distinction between phenylalanine and tryptophan moieties from a mixture on the substrates; hence, demonstrating the selectivity of the method on the nanometer scale without further need of any labeling techniques.

Total synthesis and cytotoxicity evaluation of all ochratoxin A stereoisomers

The mycotoxin ochratoxin A is a potent inhibitor of the protein biosynthesis and known to be cytotoxic in nanomolar concentrations. In order to investigate the relationship between stereochemistry and cytotoxicity of this compound, all four ochratoxin A stereoisomers have been synthesized. Using the liver cell line Hep G2, the compounds were tested for cytotoxic and apoptotic potential. It could be shown, that the l-configuration of the phenylalanine moiety of the molecule is mostly responsible for the high cytotoxicity of ochratoxin A while the stereocenter at the dihydroisocoumarine structure is of less importance.