Different Binding Modes of Small and Large Binders of GAT1

Well‐known inhibitors of the γ‐aminobutyric acid (GABA) transporter GAT1 share a common scaffold of a small cyclic amino acid linked by an alkyl chain to a moiety with two aromatic rings. Tiagabine, the only FDA‐approved GAT1 inhibitor, is a typical example. Some small amino acids such as (R)‐nipecotic acid are medium‐to‐strong binders of GAT1, but similar compounds, such as proline, are very weak binders. When substituted with 4,4‐diphenylbut‐3‐en‐1‐yl (DPB) or 4,4‐bis(3‐methylthiophen‐2‐yl)but‐3‐en‐1‐yl (BTB) groups, the resulting compounds have similar pK_i and pIC₅₀ values, even though the pure amino acids have very different values. To investigate if small amino acids and their substituted counterparts share a similar binding mode, we synthesized butyl‐, DPB‐, and BTB‐substituted derivatives of small amino acids. Supported by the results of docking studies, we propose different binding modes not only for unsubstituted und substituted, but also for strong‐ and weak‐binding amino acids. These data lead to the conclusion that following a fragment‐based approach, not pure but N‐butyl‐substituted amino acids should be used as starting points, giving a better estimate of the activity when a BTB or DPB substituent is added.     

Preparation of All Polyester‐Based Semi‐IPN Elastomers Containing Self‐Associative or Non‐Associative Guest Chains via Post‐Blending Cross‐Linking

Mechanical properties of semi‐interpenetrating polymer network (semi‐IPN) elastomers consisting of chemical networks and self‐associative/non‐associative guest chains are demonstrated. Amorphous low T_g polyesters with thiol side groups (PE‐SH) are first synthesized by melt polycondensation. PE‐SH are then converted to polyesters containing COOH side groups (PE‐COOH) and amide side groups (PE‐amide) through Michael addition reaction of thiol groups with acrylic acid and acrylamide, respectively. Homogeneous semi‐IPN elastomers are obtained by thermal cross‐linking for bulk mixtures of PE‐COOH and PE‐amide in the presence of diepoxy cross‐linkers, where COOH and epoxy groups are reacted to form chemical cross‐links while the amide units form self‐complementary hydrogen bonds. Another sample containing non‐associative chains is also prepared by using polyester with N,N‐dimethylamide units, instead of PE‐amide. Dynamic mechanical analysis reveals that guest chain incorporation systematically brings plateau modulus reduction and a unique relaxation with higher tan δ value depending on the fraction and nature of guest chains. Tensile properties are also affected by the fraction and nature of guest chains; the incorporation of hydrogen bonded chains are beneficial to enhance breaking elongation and toughness without the sacrifice of maximum stress. The knowledge found in this work will be thus beneficial for creating tough soft materials with damping applications.     

Side chain functionality dominated the chromatography of N‐protected amino acid on molecularly imprinted polymer

Molecularly imprinted polymer (MIP) with N^α‐protected amino acid as the print molecule was prepared and used as the stationary phase for the chromatographic study of molecular recognition. Particles of MIP were prepared by photopolymerization of 4‐vinylpyridine in the presence of tert‐butyloxycarbonyl‐L ‐tyrosine (Boc‐L ‐Tyr) and packed into a column for the chromatographic resolution of Boc‐L ‐Tyr and tert‐butyloxycarbonyl‐L ‐phenylalanine (Boc‐L ‐Phe). These two N^α‐protected amino acids that differ from each other in the side chain with one hydroxyl group on the benzene ring could be well separated on the MIP. A separation factor of about two was achieved by using a mixture of acetonitrile (99.5 v/v %) and acetic acid (0.5 v/v %) as the mobile phase. Results suggest that the interaction between hydroxyl group in the side chain of amino acid and pyridine in the polymer dominated the selective adsorption of print molecule on the MIP. © 2007 Wiley Periodicals, Inc. J Appl Polym Sci, 2007     

Synthesis and functionalities of poly(N‐vinylalkylamide). XIV. Polyvinylamine produced by hydrolysis of poly(N‐vinylformamide) and its functionalization

The molecular weights of poly(N‐vinylformamide) [poly(NVF)] obtained by free‐radical polymerization were expanded from being in the range of thousands to hundreds of thousands. Primary amino groups were introduced by the hydrolysis of poly(NVF) under both acidic and basic conditions. After 2 h polyvinylamine [poly(VAm)] was given at 60°C under a 2N NaOH solution. The apparent activation energy of poly(NVF) hydrolysis was 61.8 kJ/mol. Furthermore, alkyl side chains were partly introduced by a polymer modification reaction in poly(VAm) with carboxylic acid, using WSC (water‐soluble carbodiimide) as the activating agent to produce the stimuli‐responsive poly(VAm) derivative. The effects of external stimuli such as temperature and pH on the phase‐transition behavior of the copolymers were then studied. The lower critical solution temperature at pH 12 decreased depending on the alkyl group content. The phase‐transition behavior of the resulting polymers was also found to vary depending on the side‐chain length of the alkyl groups. © 2003 Wiley Periodicals, Inc. J Appl Polym Sci 89: 1277–1283, 2003     

Baceridin,a Cyclic Hexapeptide from an Epiphytic Bacillus Strain,Inhibits the Proteasome

A new cyclic hexapeptide, baceridin ( 1 ), was isolated from the culture medium of a plant‐associated Bacillus strain. The structure of 1 was elucidated by HR‐HPLC‐MS and 1D and 2D NMR experiments and confirmed by ESI MS/MS sequence analysis of the corresponding linear hexapeptide 2 . The absolute configurations of the amino acid residues were determined after derivatization by GC‐MS and Marfey's method. The cyclopeptide 1 consists partially of nonribosomal‐derived D ‐ and allo‐D ‐configured amino acids. The order of the D ‐ and L ‐leucine residues within the sequence cyclo(‐L ‐Trp‐D ‐Ala‐D ‐allo‐Ile‐L ‐Val‐D ‐Leu‐L ‐Leu‐) was assigned by total synthesis of the two possible stereoisomers. Baceridin ( 1 ) was tested for antimicrobial and cytotoxic activity and displayed moderate cytotoxicity (1–2 μg mL^?1) as well as weak activity against Staphylococcus aureus. However, it was identified to be a proteasome inhibitor that inhibits cell cycle progression and induces apoptosis in tumor cells by a p53‐independent pathway.     

Synthesis of 20‐O‐linked 20(S)‐Camptothecin Glycoconjugates: Impact of the Side Chain of the Ester‐linked Amino Acid on Epimerization During the Acylation Reaction and on Hydrolytic Stability of the Final Glycoconjugates

To improve solubility and tumor selectivity of 20(S)‐camptothecin the synthesis of 20‐O‐linked glycoconjugates 11A — G is described. Particular focus of the paper is the utilization of N‐tert‐butoxycarbonyl protected amino acid N‐carboxy anhydrides (UNCAs) 2a — f for an efficient acylation of the sterically hindered and deactivated tertiary 20‐hydroxy group of 20(S)‐camptothecin 1 . Depending on the solvent and on the side chain of the amino acid different extents of epimerization of the amino acids during the coupling reaction are observed; however, the epimers can easily be separated after removal of the tert‐butoxycarbonyl protecting group and camptothecin amino acid conjugates 4B — E with L‐ and D‐ configured amino acids, respectively, are obtained. Particularly, bulky and β‐branched amino acids can be attached to camptothecin in high yields and with low epimerization. Starting from the camptothecin amino acid conjugates 4B — E the synthesis of the glycoconjugates 11A — G is straightforward following standard procedures. The glycoconjugate hydrochlorides 11A — G show good water solubility (> 5mg‐ ml) and hydrolytic stability of the ester bond which again depends on the side chain of the amino acid residue linked to camptothecin. Particularly, glycoconjugates 11B — E with a bulky and β‐branched amino acid at the ester linkage show high hydrolytic stability in aqueous solutions with less than 2.5% of 20(S)‐camptothecin cleaved within 24 h. These results provide a basis for the selection of appropriate spacer groups for delivery systems of 20(S)‐camptothecin for therapeutic use.     

Alanine Scan of the Peptide Antibiotic Feglymycin: Assessment of Amino Acid Side Chains Contributing to Antimicrobial Activity

The antibiotic feglymycin is a linear 13‐mer peptide synthesized by the bacterium Streptomyces sp. DSM 11171. It mainly consists of the nonproteinogenic amino acids 4‐hydroxyphenylglycine and 3,5‐dihydroxyphenylglycine. An alanine scan of feglymycin was performed by solution‐phase peptide synthesis in order to assess the significance of individual amino acid side chains for biological activity. Hence, 13 peptides were synthesized from di‐ and tripeptide building blocks, and subsequently tested for antibacterial activity against Staphylococcus aureus strains. Furthermore we tested the inhibition of peptidoglycan biosynthesis enzymes MurA and MurC, which are inhibited by feglymycin. Whereas the antibacterial activity is significantly based on the three amino acids D ‐Hpg1, L ‐Hpg5, and L ‐Phe12, the inhibitory activity against MurA and MurC depends mainly on L ‐Asp13. The difference in the position dependence for antibacterial activity and enzyme inhibition suggests multiple molecular targets in the modes of action of feglymycin.     

Effective hole‐injection characteristics of organic light‐emitting diodes due to fluorinated self‐assembled monolayer embedded as a buffer layer

The electrical and optical properties of organic light‐emitting diodes (OLEDs) are demonstrated by varying the length of the alkyl chain of a fluorinated self‐assembled monolayer (F‐nSAM). OLEDs containing F‐nSAMs that have a long alkyl chain length were found to exhibit excellent properties in terms of current density, luminance, turn‐on voltage, etc. The obtained current density at 6 V, which was the highest measurement voltage, was up to about 36 times higher for an OLED including an F‐12SAM thin film with the longest chain length than for an OLED including only an indium tin oxide (ITO) substrate. With regard to luminance characteristics depending on voltage, the luminance was about 13 times higher for the OLED including the F‐12SAM thin film than for the OLED including only the ITO substrate. Also, the turn‐on voltage of the OLED including the F‐12SAM thin film was decreased by approximately 1 V compared to that of the OLED including only the ITO substrate. Although F‐nSAMs with alkyl chains have insulating film properties, F‐nSAMs with long alkyl chains exhibited good electrical and optical properties because of an improvement in the hole‐injection barrier due to a large positive shift of the vacuum level and smooth carrier injection resulting from a high contact angle due to strong hydrophobic properties caused by the good alignment properties of F‐nSAMs resulting from strong van der Waals forces between the molecules due to the long alkyl chains. © 2019 Society of Chemical Industry     

Isoprenoid Biosynthesis Inhibitors Targeting Bacterial Cell Growth

We synthesized potential inhibitors of farnesyl diphosphate synthase (FPPS), undecaprenyl diphosphate synthase (UPPS), or undecaprenyl diphosphate phosphatase (UPPP), and tested them in bacterial cell growth and enzyme inhibition assays. The most active compounds were found to be bisphosphonates with electron‐withdrawing aryl‐alkyl side chains which inhibited the growth of Gram‐negative bacteria (Acinetobacter baumannii, Klebsiella pneumoniae, Escherichia coli, and Pseudomonas aeruginosa) at ～1–4 μg mL^?1 levels. They were found to be potent inhibitors of FPPS; cell growth was partially “rescued” by the addition of farnesol or overexpression of FPPS, and there was synergistic activity with known isoprenoid biosynthesis pathway inhibitors. Lipophilic hydroxyalkyl phosphonic acids inhibited UPPS and UPPP at micromolar levels; they were active (～2–6 μg mL^?1) against Gram‐positive but not Gram‐negative organisms, and again exhibited synergistic activity with cell wall biosynthesis inhibitors, but only indifferent effects with other inhibitors. The results are of interest because they describe novel inhibitors of FPPS, UPPS, and UPPP with cell growth inhibitory activities as low as ～1–2 μg mL^?1.     

Identification and Characterization of Phospholipids with Very Long Chain Fatty Acids in Brewer’s Yeast

Yeast lipids and fatty acids (FA) were analyzed in Saccharomyces pastorianus from seven breweries and in the dietary yeast supplement Pangamin. GC–MS identified more than 30 FA, half of which were very‐long chain fatty acids (VLCFA) with hydrocarbon chain lengths of ≥22 C atoms. Positional isomers ω‐9 and ω‐7 were identified in FA with C18–C28 even‐numbered alkyl chains. The most abundant ω‐7 isomer was cis‐vaccenic acid. The structure of monounsaturated FA was proved by dimethyl disulfide adducts (position of double bonds and cis geometric configuration) and by GC–MS of pyridyl carbinol esters. Ultra‐high performance liquid chromatography‐tandem mass spectrometry with negative electrospray ionization identified the phospholipids phosphatidylethanolamine, phosphatidylinositol and phosphatidylcholine, with more than 150 molecular species. Wild‐type unmutated brewer's yeast strains conventionally used for the manufacture of food supplements were found to contain VLCFA.     

Nucleophilic Water Capture or Proton Loss: Single Amino Acid Switch Converts δ‐Cadinene Synthase into Germacradien‐4‐ol Synthase

δ‐Cadinene synthase is a sesquiterpene cyclase that utilises the universal achiral precursor farnesyl diphosphate (FDP) to generate predominantly the bicyclic sesquiterpene δ‐cadinene and about 2 % germacradien‐4‐ol, which is also generated from FDP by the cyclase germacradien‐4‐ol synthase. Herein, the mechanism by which sesquiterpene synthases discriminate between deprotonation and reaction with a nucleophilic water molecule was investigated by site‐directed mutagenesis of δ‐cadinene synthase. If W279 in δ‐cadinene synthase was replaced with various smaller amino acids, the ratio of alcohol versus hydrocarbon product was directly proportional to the van der Waals volume of the amino acid side chain. DCS‐W279A is a catalytically highly efficient germacradien‐4‐ol synthase (k_cat/K_M=1.4×10^?3 μm s^?1) that produces predominantly germacradien‐4‐ol in addition to 11 % δ‐cadinene. Water capture is not achieved through strategic positioning of a water molecule in the active site, but through a coordinated series of loop movements that allow bulk water access to the final carbocation in the active site prior to product release.     

Carbon assimilation and dissimilation during growth of Geotrichum candidum on amino acids and glucose

BACKGROUND: This work examines the metabolic behaviour of amino acids during Geotrichum candidum growth, in the presence of a primary carbon source like glucose. Amino acids were characterized based on their carbon assimilation and dissimilation by G. candidum, in the presence of glucose as the limiting substrate. RESULTS: The first group (Cys, His, Phe, Thr and Trp) was only used as nitrogen sources by G. candidum, with glucose being the carbon and energy source. Glucose repression was shown for the rest of the amino acids, since only after glucose depletion amino acids from the second group (Gly, Lys, Met, Val) were dissimilated for energy supply by oxidation into CO₂, while those from the third group (Ala, Arg, Asp, Glu, Leu, Pro and Ser) were assimilated as carbon sources (and additionally used as nitrogen sources), leading to a diauxic growth. CONCLUSION: This energy‐saving response was not previously shown for the second fungus involved in ripening of soft white cheese—P. camembertii—leading to simultaneous use of some amino acids and glucose as carbon and energy sources, and hence lower growth rates than those recorded during G. candidum growth. Copyright © 2007 Society of Chemical Industry     

Alternative molecularly imprinted membranes from a derivative of natural polymer,cellulose acetate

Molecularly imprinted polymeric membranes were prepared from cellulose acetate (CA), of which acetyl content was 40%, by applying the alternative molecular imprinting technique. The Z‐d ‐Glu imprinted polymeric membranes thus obtained recognized d ‐Glu in preference to l ‐Glu from racemic Glu mixtures and vice versa. The affinity constants between Glu and the chiral recognition site for two kinds of membranes were determined to be 3.1 × 10³ mol⁻¹ dm³ from the adsorption isotherm of d ‐Glu or L ‐Glu in the molecularly imprinted CA membranes. Enantioselective electrodialysis was attained with the present membranes reflecting their adsorption selectivity. d ‐Glu was preferentially permeated through the Z‐d ‐Glu imprinted CA membrane, whereas L ‐Glu was permeated through the Z‐L ‐Glu imprinted CA membrane. The present study suggests that the molecularly imprinted CA membranes are applicable to the optical resolution of racemic amino acids. © 1999 John Wiley & Sons, Inc. J Appl Polym Sci 72: 493–499, 1999     

Synthesis and characterization of biodegradable block copolymer pluronic‐b‐poly(L‐lysine)

This study presented the investigations on the synthesis of a novel biodegradable block copolymer of pluronic‐b‐poly(L ‐lysine) (pluronic‐b‐PLL), which combined the characteristics of aliphatic polyester and poly(amino acids). The synthesis work started with end‐capping of pluronic with N‐t‐butoxycarbonyl‐L ‐phenylalanine using dicyclohexylcarbodiimide in the presence of 4‐dimethylaminopyridine, followed by a deprotection process to obtain the amino‐terminated pluronic; the new primary amino group in the modified pluronic initiated ring‐opening polymerization of amino acid N‐carboxyanhydride, which afforded the pluronic‐b‐poly(N^ε‐(Z)‐L ‐lysine) block copolymer. Finally, removal of the side‐chain N^ε‐(carbonybenzoxy) end protecting groups yields the block copolymer of pluronic‐b‐PLL. The products were characterized by ¹H‐NMR, FTIR, DSC, and GPC. The block copolymer micelle containing the anticancer drug paclitaxel was prepared by the double emulsion method. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2009     

Jamming rheology of model cementitious suspensions composed of comb‐polymer stabilized magnesium oxide particles

The colloidal microstructure of concentrated suspensions containing anionic comb‐polymer‐stabilized magnesium oxide (MgO) particles in water was analyzed by shear rheometry for indications of changes in particle microstructure based on particle size and comb‐polymer usage. As the suspensions were sheared at different rates, jamming in the sheared MgO suspensions was observed as shear stress overshoots. The shear‐induced evolution of the suspension's microstructure was strongly related to the perceived interactions between neighboring MgO particles in the suspension. In the jammed state, interactions are believed to be enhanced by the formation of entanglements between opposing comb‐polymer side‐chains. Steric repulsion between side‐chains was lessened for large particles on account of their diameters, which further enabled side‐chain entanglement during close particle contact under shear. Suspensions with relatively wide particle size distributions (0.5–400 μm) were theorized to form hydrocluster aggregates, while suspensions with narrower particle size distributions (0.5–40 μm) most likely resulted in networked microstructures under the influence of the chain entanglements from the adsorbed comb‐polymer. © 2014 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014 , 131, 40429.     

Stabilization of a monoclonal antibody during purification and formulation by addition of basic amino acid excipients

BACKGROUND: Protein stability of therapeutic monoclonal antibodies during processing and final storage is imperative for the commercial success of the product. Amino acid addition is one of the options for stabilization of proteins that can be employed during the manufacturing process and storage. RESULTS: Use of arginine in the elution buffer during Protein A purification and subsequent neutralization doubled the yield of antibody compared with the original glycine‐based elution buffer. The role of amino acids in stabilizing monoclonal antibody liquid formulations was then studied using differential scanning calorimetry. Increase in the unfolding transition temperatures (T_m) of immunoglobulin G (IgG) was measured after supplementing a glycine buffer with a range of amino acids. The basic amino acids histidine, lysine, and arginine stabilized all three domains of IgG. The neutral amino acids serine and alanine provided less stabilization; glutamine, proline, and the acidic amino acids provided negligible stabilization. CONCLUSION: The positive charge on the side‐chain of histidine, lysine, and arginine appears to be the most important factor affecting the IgG stabilization. It is probable that the mechanism for IgG stabilisation is the same for all of the basic amino acids and that it binds transiently to IgG side chains, altering water populations in the solvation shell, making unfolding and aggregation less energetically favourable. Copyright © 2011 Society of Chemical Industry     

Optical resolution of racemic amino acid derivatives with chiral polyamides bearing glutamyl residue as a diacid component

Novel chiral polyamides with chiral environment in their main chains were obtained from aromatic diamine, 4,4′‐diaminodiphenylmethane (DADPM), and the D ‐isomer or the L ‐isomer of N‐α‐protected glutamic acid, such as N‐α‐benzyloxycarbonyl‐D ‐glutamic acid (Z‐D ‐Glu‐OH) or N‐α‐benzyloxycarbonyl‐L ‐glutamic acid (Z‐L ‐Glu‐OH), in the presence of triphenyl phosphite (TPP). Two types of newly prepared polyamide showed optical rotation, implying that there were asymmetric carbons in their main chains. Circular dichroism studies demonstrated that resulting chiral poly‐ amides took a helical structure. Optical resolution ability of those two types of polyamide was studied by adopting potential difference as a driving force for membrane transport. Membranes showed permselectivity toward racemic mixture of N‐α‐acetyltryptophan (Ac‐Trp). The permselectivity was dependent on the absolute configuration of diacid component. The permselectivity was expressed by diffusivity selectivity, which was determined by the presence of chiral helicity. © 2011 Wiley Periodicals, Inc. J Appl Polym Sci, 2011     

Synthesis and antimicrobial activity of some cross‐linked copolymers with alkyl chains of various lengths

Amphoteric polymer hydrogels were prepared by the copolymerization of three kinds of N,N′‐dimethyl‐N‐alkylmethacryloxylethyl ammoniumbromide (DMAEA) with different lengths of alkyl chains (DMAEA‐RB) (R‐ehtyl/hexyl/dodecyl), acrylic acid (AA), and acrylamide (AM). The water content of the AA‐AM‐(DMAEA‐RB) terpolymers decreased with the increasing length of alkyl chains in quaternary ammonium group in the terpolymers and increased with the increasing molar ratio of DMAEA‐RB to 2 mol % and then decreased. Their antibacterial activities against Escherichia coli and Staphylococcus hyicus were investigated by a colony count method. It was found that the copolymer exhibited higher antibacterial activity with increasing chain length of alkyl groups in ammonium groups. For P[AA‐AM‐(DMAEA‐DB)], the DMAEA‐DB content is higher and contact time is longer, its antibacterial activity is better. However, when the contact time and quaternary ammonium content were above 30 min and 2%, respectively, the amount of live cells N(t) in a cell suspension increased in the presence of P[AA‐AM‐(DMAEA‐EB)] or P[AA‐AM‐(DMAEA‐HB)]. © 2010 Wiley Periodicals, Inc. J Appl Polym Sci, 2011     

Synthesis and Surface-Active Properties of a Homologous Series of Star-Like Triple-Chain Anionic Surfactants Derived from 1,1,1-Tris(hydroxymethyl)ethane

A novel homologous series of trimeric anionic surfactants, 3C_nTE3CNa (where n is a fatty acid chain length of 7, 10, or 12), with three hydrocarbon chains and three carboxylate heads connected via tri‐etheric bonds were synthesized from long‐chain α‐bromo fatty acids and a triol, 1,1,1‐tris(hydroxymethyl)ethane. The obtained trimeric carboxylic acids were esterified and purified by silica gel column chromatography, then hydrolyzed with dilute sodium hydroxide solution to form a series of trimeric carboxylate surfactant products. All prepared compounds were analyzed by IR, ¹H NMR, and ¹³C NMR spectroscopy to confirm their chemical structures. Their surface‐active properties were investigated. The critical micelle concentrations (cmc) of 3C_nTE3CNa were in the range of 0.12–0.71 mmol/L, and the surface tensions at the cmc (γ_cmc) were 29.3–34.8 mN/m.     

Surface characteristics of poly(γ‐alkyl α l‐glutamate)s with different alkyl groups

A series of poly(γ‐alkyl α L ‐glutamate)s with different alkyl groups were synthesized by the ring opening polymerization of corresponding α‐amino acid N‐carboxyanhydrides. The characteristics of these polyglutamate surfaces were evaluated by attenuated total reflectance Fourier transform infrared spectroscopy spectra, water contact angle, water absorption, protein adsorption, and platelet adhesion measurements. Changing the length of the alkyl side chain provides a unique opportunity to study the influence of carbon number in the alkyl group on the surface properties of the polyglutamates. Water contact angle and water absorption data show that the hydrophilicity of these polyglutamate surfaces decreases with the increasing of methylene in the alkyl group. Protein adsorption on these polyglutamate surfaces increases with the enhancing of surface hydrophobicity. However, the changes in platelets adhesion could be attributed to the hydrophilicity/hydrophobicity of the polyglutamates and the specific effect of alkyl group. © 2010 Wiley Periodicals, Inc. J Appl Polym Sci, 2011