Synthesis and electrochemical behaviour of β-halodehydroamino acid derivatives

Several new β,β-dihalo and β-halo-β-substituted dehydroalanines and dehydrodipeptides were synthesized by reacting the corresponding dehydroamino acid derivative with a N-halosuccinimide or in the case of β,β-di-iododehydroalanines with iodine. The results obtained confirmed that the stereochemical outcome of the halogenation reaction with β-substituted dehydroamino acids depends on the substrate. Thus, an increase Z-stereoselectivity was found when the β-phenyldehydroalanines were used as substrates and when these compounds were N-protected with 4-tolylsulfonyl or with carbamates. From this study, it is also possible to conclude that when N-iodosuccinimide was used as reagent a much higher Z-stereoselectivity is found. The electrochemical behaviour of the halogenated dehydroamino acids was studied by cyclic voltammetry. The results show a shift in the reduction peak to higher potentials of the β-halogenated dehydroamino acids when compared with the corresponding non-halogenated derivatives. As expected, the β,β-dihalodehydroalanines exhibit higher peak potentials than β-halo-β-substituted dehydroalanines and the bromo derivatives have lower peak potentials when compared with the corresponding iododehydroamino acids. Controlled potential electrolysis of several β-halo-β-substituted dehydroamino acids afforded the corresponding dehalogenated dehydroamino acids as mixtures of their E and Z-isomers. In all cases, the major isomer isolated results from dehalogenation without isomerization. These new results show that electrochemical reduction constitutes a valuable method for the synthesis of the E-isomer of β-substituted dehydroalanines.     

Clavulanic acid, a β-lactamase inhibitor: biosynthesis and molecular genetics

Clavulanic acid is a secondary metabolite produced by Streptomyces clavuligerus. It possesses a clavam structure and a characteristic 3R,5R stereochemistry essential for action as a β-lactamase inhibitory molecule. It is produced from glyceraldehyde-3-phosphate and arginine in an eight step biosynthetic pathway. The pathway is carried out by unusual enzymes, such as (1) the enzyme condensing both precursors, N ²-(2-carboxyethyl)-arginine (CEA) synthetase, (2) the β-lactam synthetase cyclizing CEA and (3) the clavaminate synthetase, a well-characterized multifunctional enzyme. Genes for biosynthesis of clavulanic acid and other clavams have been cloned and characterized. They offer new possibilities for modification of the pathway and for obtaining new molecules with a clavam structure. The state of the regulatory proteins controlling clavulanic acid biosynthesis, as well as the relationship between the biosynthetic pathway of clavulanic acid and other clavams, is discussed. Received: 9 February 2000 / Received revision: 10 May 2000 / Accepted: 12 May 2000     

Studies on amino acid replacement and inhibitory activity of a β-lactamase inhibitory peptide

An SHV β-lactamase gene was amplified from a β-lactam resistant Klebsiella pneumoniae K-71 genomic DNA. After expression and purification, we demonstrated that peptide P1 could inhibit the hydrolysis activity of both TEM-1 and SHV β-lactamase in vitro. Three mutations were introduced into P1 in which the first residue S was replaced by F, the 18th residue V was mutated to Y, and the 15th residue Y was substituted with A, C, G, and R to obtain the mutants of P1-A, P1- C, P1-G, and P1-R, respectively. The mutant peptides were purified and their inhibitory constants against TEM-1 and SHV β-lactamase were determined. All these β-lactamase inhibitory peptides could inhibit the activity of both β-lactamases, while the mutant peptides showed stronger inhibitory activities against TEM-1 β-lactamase than against SHV β-lactamase. Inhibition data suggested that P1-A improved the β-lactamase inhibitory activity by over 3-fold compare to P1. When P1-A was incubated with K. pneumoniae K-71 in Luria-Bertani medium containing ampicillin, it showed a much stronger growth of inhibition ratio over P1. This study gives us a good candidate for development of novel β-lactamase inhibitors.     

Synthesis of β-galactosyl-(hydroxy amino acid) derivatives using β-galactosidase activity ofAchatina achatina digestive juice

Summary The transglycosylation activity of β-galactosidase fromAchatina achatina digestive juice was tested for glycosylating protected hydroxy amino acids. Attractive yields of β-galactosyl-(Z-Ser-OMe) (35%) and β-galactosyl-(Z-Hyp-OMe) (28%) could be obtained using lactose as glycosyl donor and the corresponding amino acid methyl esters N-protected by a benzyloxycarbonyl group (Z) as glycosyl acceptors.     

Synthesis,biological evaluation,and structure–activity relationship study of novel cytotoxic aza-caffeic acid derivatives

Three series of aza-caffeic acid derivatives with different linkers were designed and synthesized. Each of the synthesized derivatives was then used in cytotoxicity screening on either 8 or 12 human cancer cell lines. The structure–activity relationships on three structural regions A, B, and C are analyzed in detail, indicating that a nine bond linker B, containing a piperazine unit, is the most favorable linker leading to the generation of molecules with potent cytotoxicities. Compound (E)-1-(4-(3,4-dichlorobenzyl)piperazin-1-yl)-3-(4-(4-ethoxybenzyloxy)-3,5-dimethoxyphenyl)prop-2-en-1-one (80) exhibited the most significant and selective cytotoxicity to KB, BEL7404, K562, and Eca109 cell lines, with IC₅₀ values of 0.2, 2.0, 1.7, and 1.1 μM, respectively, stronger than that seen for caffeic acid phenethyl ester (CAPE) and cisplatin (CDDP). Flow cytometric and western blot analysis indicate that compound 80 plays a role in mitochondria-dependent apoptosis activity by suppressing K562 cell proliferation in a concentration- and time-dependent manner.     

Kojic acid derived hydroxypyridinone–chloroquine hybrids: Synthesis,crystal structure,antiplasmodial activity and β-haematin inhibition

Aminochloroquinoline–kojic acid hybrids were synthesized and evaluated for β-haematin inhibition and antiplasmodial activity against drug resistant (K1) and sensitive (3D7) strains of Plasmodium falciparum. Compound 7j was the most potent compound in both strains (IC₅₀^3D7 = 0.004 μM; IC₅₀^K1 = 0.03 μM) and had the best β-haematin inhibition activity (0.07 IC₅₀ equiv vs 1.91 IC₅₀ equiv for chloroquine). One compound 8c was found to be equipotent in both strains (IC₅₀ = 0.04 μM).     

Design,synthesis, and structure–activity relationship of novel and effective apixaban derivatives as FXa inhibitors containing 1,2,4-triazole/pyrrole derivatives as P2 binding element

Four series of novel and potent FXa inhibitors possessing the 1,2,4-triazole moiety and pyrrole moiety as P2 binding element and dihydroimidazole/tetrahydropyrimidine groups as P4 binding element were designed, synthesized, and evaluated for their anticoagulant activity in human and rabbit plasma in vitro. Most compounds showed moderate to excellent activity. Compounds 14a, 16, 18c, 26c, 35a, and 35b were further examined for their inhibition activity against human FXa in vitro and rat venous thrombosis in vivo. The most promising compound 14a, with an IC₅₀ (FXa) value of 0.15 μM and 99% inhibition rate, was identified for further evaluation as an FXa inhibitor.     

Synthesis of disaccharide derivatives employing β-N-acetyl- d-hexosaminidase, β-d-galactosidase and β-d-glucuronidase

-N-Acetyl-d-hexosaminidase from Aspergillus oryzae catalysed the stereo- and regiospecific formation of the 6-O-benzylated disaccharide derivatives GalNAc1-3(6- OBn)Gal-SEt and GlcNAc1-3(6-OBn)Gal-SEt, which were obtained in transglycosylation reactions employing ethyl 6- O-benzyl-1-thio--d-galactopyranoside as acceptor. Preparative amounts of the chitobiose derivative GlcNAc1- 3GlcNAc-OPhNO2-p was prepared as well. - N-Acetyl-d-hexosaminidase from bovine testes catalysed the specific synthesis of GlcNAc1-3(6-OBn)GlcNH2-SEt and GalNAc1-3(6-OBn)GlcNH2-SEt, employing ethyl 2-amino-6-O-benzyl-2-deoxy-1-thio--d-glucopyranoside as acceptor. -d-Glucuronidase from E. coli was found to catalyse the formation of GlcA1-3(6-OBn)GlcNH2- SEt employing the same acceptor.     

Twelve positions in a β-lactamase that can expand its substrate spectrum with a single amino acid substitution

The continuous evolution of β-lactamases resulting in bacterial resistance to β-lactam antibiotics is a major concern in public health, and yet the underlying molecular basis or the pattern of such evolution is largely unknown. We investigated the mechanics of the substrate fspectrum expansion of the class A β-lactamase using PenA of Burkholderia thailandensis as a model. By analyzing 516 mutated enzymes that acquired the ceftazidime-hydrolyzing activity, we found twelve positions with single amino acid substitutions (altogether twenty-nine different substitutions), co-localized at the active-site pocket area. The ceftazidime MIC (minimum inhibitory concentration) levels and the relative frequency in the occurrence of substitutions did not correlate well with each other, and the latter appeared be largely influenced by the intrinsic mutational biases present in bacteria. Simulation studies suggested that all substitutions caused a congruent effect, expanding the space in a conserved structure called the omega loop, which in turn increased flexibility at the active site. A second phase of selection, in which the mutants were placed under increased antibiotic pressure, did not result in a second mutation in the coding region, but a mutation that increased gene expression arose in the promoter. This result suggests that the twelve amino acid positions and their specific substitutions in PenA may represent a comprehensive repertoire of the enzyme's adaptability to a new substrate. These mapped substitutions represent a comprehensive set of general mechanical paths to substrate spectrum expansion in class A β-lactamases that all share a functional evolutionary mechanism using common conserved residues.     

Ruthenium complexes endowed with potent anti-Trypanosoma cruzi activity: Synthesis,biological characterization and structure–activity relationships

Although effective against epimastigotes (proliferative form) and of low cytotoxicity in mammals, the aryl-4-oxothiazolylhydrazones (ATZ) display only limited activity against trypomastigotes (bloodstream form) of Trypanosoma cruzi. Considering the metal complexation approach with bioactive ligands as one possible strategy for improving the biological efficacy of ATZ, a set of eight new ruthenium–ATZ complexes (RuCl₂ATZCOD, COD is 1,5-cyclooctadiene) were prepared, chemically and biologically characterized, including in vitro assays against epimastigotes and trypomastigote forms of the parasite and also assessment of cytotoxicity in mammals. Two of these complexes presented antitrypanosomal activity at non-cytotoxic concentrations on mammalian cells and of higher potency than its metal–free ligands, while the metallic precursor [RuCl₂COD(MeCN)₂] showed only moderate antitrypanosomal activity. Comparative analysis between the ruthenium complexes and metal–free ligands demonstrated the usefulness of this approach, with the establishment of new SAR data. Additional pharmacological tests, including a DNA bond assay, gave rise to the proposal of a single preliminary explanation for the molecular origin of the bioactivity.     

Synthesis and human carbonic anhydrase I,II, VA,and XII inhibition with novel amino acid–sulphonamide conjugates

Abstract

A series of amino acid–sulphonamide conjugates was prepared through benzotriazole mediated coupling reactions and characterised by ¹H-NMR, ¹³C-NMR, MS, and FTIR spectroscopic techniques as well as elemental analysis. The carbonic anhydrase (CA, EC 4.2.1.1) inhibitory activity of the new compounds was determined against four human (h) isoforms, hCA I, hCA II, hCA VA, and hCA XII. Most of the synthesised compounds showed effective in vitro CA inhibitory properties. The new amino acid–sulphonamide conjugates showed potent inhibitory activity against hCA II, some of them at subnanomolar levels, exhibiting more effective inhibitory activity compared to the standard drug acetazolamide. Some of these sulphonamides were also found to be effective inhibitors of hCA I, hCA VA, and hCA XII, with activity from the low to high nanomolar range.     

Relationship between structure and amino acid sequence of strongly twisted and coiled β-hairpins in globular proteins

β-Hairpins are widespread in proteins, and it is possible to find them both within β-sheets and separately. In this work, a comparative analysis of amino acid sequences of β-strands within strongly twisted β-hairpins from different structural protein subclasses has been conducted. Strongly twisted and coiled β-hairpin generates in the space a right double helix out of β-strands that are connected by a loop region (connections). The frequencies of amino acid residues on the internal (concave) and external (convex) surfaces of strongly twisted β-hairpins have been determined (220 β-hairpins from nonhomologous proteins were studied). The concave surface of these β-hairpins is mainly generated by hydrophobic residues, while the convex surface by hydrophilic residues; accordingly, the alternation of hydrophobic internal and hydrophilic external residues is observed in their amino acid sequences. Amino acid residues of glycine and alanine (especially in places of the largest twisting of the strands) were anomalously frequently found in internal positions of strongly twisted and coiled β-hairpins. It was established that internal positions never contain the proline residues, while external positions in the twisting region contain them in a relatively large amount. It was demonstrated that at least one amino acid residue in α_L- or ε-conformation is required for generation of relatively short (up to 7 amino acid residues) connection. As a rule, these positions are occupied by glycines. Thus, not only the alternation of hydrophobic and hydrophilic amino acid residues, but also the presence of one or two glycine residues in the connection region and the excess of glycines and alanines in the places of the largest strand twisting on the concave surface, as well as the presence of prolines on the convex surface, are required to generate a strongly twisted and coiled β-hairpin.     

Synthesis and structure–activity relationship of thiobarbituric acid derivatives as potent inhibitors of urease

A series of thiobarbituric acid derivatives 1–27 were synthesized and evaluated for their urease inhibitory potential. Exciting results were obtained from the screening of these compounds 1–27. Compounds 5, 7, 8, 11, 16, 17, 22, 23 and 24 showed excellent urease inhibition with IC₅₀ values 18.1 ± 0.52, 16.0 ± 0.45, 16.0 ± 0.22, 14.3 ± 0.27, 6.7 ± 0.27, 10.6 ± 0.17, 19.2 ± 0.29, 18.2 ± 0.76 and 1.61 ± 0.18 μM, respectively, much better than the standard urease inhibitor thiourea (IC₅₀ = 21 ± 0.11 μM). Compound 3, 4, 10, and 26 exhibited comparable activities to the standard with IC₅₀ values 21.4 ± 1.04 and 21.5 ± 0.61μM, 22.8 ± 0.32, 25.2 ± 0.63, respectively. However the remaining compounds also showed prominent inhibitory potential The structure–activity relationship was established for these compounds. This study identified a novel class of urease inhibitors. The structures of all compounds were confirmed through spectroscopic techniques such as EI-MS and ¹H NMR.     

Synthesis and anti-TMV activity of novel β-amino acid ester derivatives containing quinazoline and benzothiazole moieties

Here, a series of β-amino acid ester derivatives containing quinazoline and benzothiazoles was synthesized and evaluated for anti-tobacco mosaic virus (TMV) activity. The compounds 3n, 3o, 3p and 3q showed good antiviral activity against TMV at a concentration of 500 μg/mL, with curative rates of 55.55%, 52.32%, 52.77% and 50.91%, respectively, and protection rates of 52.33%, 55.96%, 54.21% and 50.98%, respectively. These values were close to those of the commercially available antiviral agent ningnanmycin (which has curative and protection rates of 55.27% and 52.16%, respectively). To our knowledge, this is the first report of the anti-TMV activity of β-amino acid ester derivatives containing quinazoline and benzothiazoles moieties; the results indicate that these novel compounds can potentially be used as protective agents against TMV diseases.     

Effects of the amino acid derivatives, β-hydroxy-β-methylbutyrate,taurine, and N-methyltyramine,on triacylglycerol breakdown in fat cells

Journal of Physiology and Biochemistry - Various amino acid (AA) metabolites are used as supplements to facilitate metabolic control and enhance responsiveness of insulin-sensitive tissues....     

Synthesis of chimera oligopeptide including furanoid β-sugar amino acid derivatives with free OHs: mild but successful removal of the 1,2-O-isopropylidene from the building block

Complementary to hydrophobic five membered ring β-amino acids (e.g. ACPC), β-sugar amino acids (β-SAAs) have found increasing application as hydrophilic building blocks of foldamers and α/β chimeric peptides. Fmoc-protected β-SAAs [e.g. Fmoc-RibAFU(ip)-OH] are indeed useful Lego elements, ready to use for SPPS. The removal of 1,2-OH isopropylidene protecting group increasing the hydrophilicity of such SAA is presented here. We first used N₃-RibAFU(ip)-OH model compound to optimize mild deprotection conditions. The formation of the 1,2-OH free product N₃-RibAFU-OH and its methyl glycoside methyl ester, N₃-RibAFU(Me)-OMe were monitored by RP-HPLC and found that either 50% TFA or 8 eqv. Amberlite IR-120 H⁺ resin in MeOH are optimal reagents for the effective deprotection. These conditions were then successfully applied for the synthesis of chimeric oligopeptide: -GG-X-GG- [X=RibAFU(ip)]. We found the established conditions to be effective and—at the same time—sufficiently mild to remove 1,2-O-isopropylidene protection and thus, it is proposed to be used in the synthesis of oligo- and polypeptides of complex sequence combination.

     

Synthesis, crystallography, phosphorescence of platinum complexes coordinated with 2-phenylpyridine and a series of β-diketones

Metallocyclic platinum(II) complexes coordinating with 2-phenylpyridine (ppy) and a series of β-diketone ancillary O^∧O ligands, (ppy)Pt(acac), (ppy)Pt(ba), (ppy)Pt(dbm), and (ppy)Pt(tta) (acac = acetylacetone, ba = benzoylacetone, dbm = dibenzoylmethane, tta = thenoyltrifluoroacetone) were synthesized. The crystal structure, absorption, emission, quantum yield and phosphorescence life time were characterized. As the conjugative π system of the O^∧O ligand increases in the order acac < ba < dbm, or there is a group -CF₃to attract the electron density of the tta ligand, the quantum yield decreases in the order (ppy)Pt(acac) > (ppy)Pt(ba) > (ppy)Pt(dbm) > (ppy)Pt(tta) due to an energy back-flow from ppy to the O^∧O ligand, a trend also in contrast to the phosphorescence emission spectra and time decay (biexponentially, ∼0.7-13 μs).     

Synthesis, crystal structure and electrochemistry of tetrahedral mono-β-diketonato titanocenyl complexes

The synthesis of a variety of tetrahedral β-diketonato titanium(IV) complexes of the type with R = CF₃, OCH₃, C₆H₅, CH₃ and Fc is described. The Ti^III/Ti^IV couples and the Fc/Fc⁺ couple exhibited chemically and electrochemically reversible cyclic voltammetric behaviour. The formal reduction potential of the Ti^III/IV couple increased as the group electronegativity of the R group of the β-diketonato ligand increased. Bulk electrolysis showed that one electron was transferred in the Ti^III/IV couple and one electron in the ferrocenyl/ferrocenium redox couple in the ligand. The crystal structure for the R = OCH₃ complex showed that this β-keto-ester binds through the carbonyl oxygen of the ester group and not the ether oxygen.     

Candida albicans mycelial wall structure: supramolecular complexes released by Zymolyase,chitinase and β-mercaptoethanol

Different techniques released from the wall of Candida albicans mycelial cells high molecular weight mannoprotein materials with different levels of complexity. SDS solubilized among others one protein of 180 kDa which reacted with a monoclonal antibody (MAb) specific of a O-glycosylated protein secreted by regenerating mycelial protoplasts [Elorza et al. (1989) Biochem Biophys Res Commun 162:1118–1125]. Zymolyase, chitinase and -mercaptoethanol, released different types of high molecular highly polydisperse mannoprotein materials (>180 kDa) that also reacted with the same MAb. These materials had N-glycosidically linked sugar chains, in addition to the O-glycosidically bonded sugars, as their molecular masses were significantly reduced by Endo H digestion. Besides, the specific materials released by either zymolyase or chitinase seemed to be the same throughout the process of germ tube formation. Transmission electron microscopy of thin sections of cells and walls showed that mannoproteins and chitin are evenly distributed throughout the entire cell wall structure.     

Melatonin analogue new indole hydrazide/hydrazone derivatives with antioxidant behavior: Synthesis and structure–activity relationships

Melatonin (MLT) is a hormone produced in the brain by the pineal gland, from the amino acid tryptophan. It is also an antioxidant hormone with a particular role in the protection of nuclear and mitochondrial DNA. In recent years, many physiological properties of MLT have been described resulting in much attention in the development of synthetic compounds possessing the indole ring. Sixteen MLT analogue indole hydrazide/hydrazone derivatives were synthesized and in vitro antioxidant activity was investigated. Most of the compounds showed significantly higher activity than MLT at 10^{? 3} M and 10^{? 4} M concentrations.