Blue fluorescent dye-protein complexes based on fluorogenic cyanine dyes and single chain antibody fragments

Fluoromodules are complexes formed upon the noncovalent binding of a fluorogenic dye to its cognate biomolecular partner, which significantly enhances the fluorescence quantum yield of the dye. Previously, several single-chain, variable fragment (scFv) antibodies were selected from a yeast cell surface-displayed library that activated fluorescence from a family of unsymmetrical cyanine dyes covering much of the visible and near-IR spectrum. The current work expands our repertoire of genetically encodable scFv-dye pairs by selecting and characterizing a group of scFvs that activate fluorogenic violet-absorbing, blue-fluorescing cyanine dyes, based on oxazole and thiazole heterocycles. The dye binds to both yeast cell surface-displayed and soluble scFvs with low nanomolar K(d) values. These dye-protein fluoromodules exhibit high quantum yields, approaching unity for the brightest system. The promiscuity of these scFvs with other fluorogenic cyanine dyes was also examined. Fluorescence microscopy demonstrates that the yeast cell surface-displayed scFvs can be used for multicolor imaging. The prevalence of 405 nm lasers on confocal imaging and flow cytometry systems make these new reagents potentially valuable for cell biological studies.     

Microwave assisted Suzuki reaction in N-butylpyridinium salts/water systems

A solvent mixture containing the ionic liquid derived from 4-picoline (C₄MPyBF₄) and water was tested for the Suzuki-Miyaura reaction under microwave irradiation. This solvent system led to very high conversions and clean reactions. When compared with the other more common ionic liquid, such as C₄MImBF₄, C₄MPyBF₄ proved to be superior.     

Contact angle dependence of the resonant frequency of sessile water droplets

The resonant vibrations of small (microliter) sessile water droplets supported on solid substrates were monitored using a simple optical detection technique. A small puff of air was used to apply an impulse to the droplets and their time dependent oscillations were monitored by passing a laser beam through the droplet and measuring the variations of the intensity of the scattered light using a simple photodiode arrangement. The resulting time dependent intensity changes were then Fourier transformed to obtain information about the vibrational frequencies of the droplets. The resonant frequencies of droplets with masses in the range 0.005-0.03 g were obtained on surfaces with water contact angles ranging from 12 ± 4° to 160 ± 5°. The contact angle dependence of the resonant frequency of the droplets was found to be in good agreement with a simple theory which considers standing wave states along the meridian profile length of the droplets.     

Novel anti-Plasmodial hits identified by virtual screening of the ZINC database

Increased resistance of Plasmodium falciparum to most available drugs challenges the control of malaria. Studies with protease inhibitors have suggested important roles for the falcipain family of cysteine proteases. These enzymes act in concert with other proteases to hydrolyze host erythrocyte hemoglobin in the parasite food vacuole. In order to find potential new antimalarial drugs, we screened in silico the ZINC database using two different protocols involving structure- and ligand-based methodologies. Our search identified 19 novel low micromolar inhibitors of cultured chloroquine resistant P. falciparum. The most active compound presented an IC₅₀ value of 0.5 μM against cultured parasites and it also inhibited the cysteine protease falcipain-2 (IC₅₀ = 25.5 μM). These results identify novel classes of antimalarials that are structurally different from those currently in use and which can be further derivatized to deliver leads suitable for optimisation.     

The synthesis,structural characterization and biological evaluation of N‐(ferrocenylmethyl amino acid) fluorinated benzene‐carboxamide derivatives as potential anticancer agents

A series of N‐(ferrocenylmethyl amino acid) fluorinated benzene‐carboxamide derivatives 4b , 4c , 4d , 4e , 4f , 4g , 4h , 4i and 5b , 5c , 5d , 5e , 5f , 5g , 5h , 5i have been synthesized by coupling ferrocenylmethyl amine 3 with various substituted N‐(fluorobenzoyl) amino acid derivatives using the standard N‐(3‐dimethylaminopropyl)‐N′‐ethylcarbodiimide hydrochloride, 1‐hydroxybenzotriazole protocol. The amino acids employed in this study were glycine and L‐alanine. All of the compounds were fully characterized using a combination of ¹H NMR, ¹³C NMR, ¹⁹F NMR, distortionless enhancement by polarization transfer (DEPT)‐135, ¹H–¹H correlation spectroscopy (COSY) and ¹H–¹³C COSY (heteronuclear multiple‐quantum correlation) spectroscopy. The compounds were biologically evaluated on the oestrogen‐positive MCF‐7 breast cancer cell line. Compounds 4g , 4i , 5h and 5i exhibited cytotoxic effects on the MCF‐7 breast cancer cell line. N‐(Ferrocenylmethyl‐L‐alanine)‐3,4,5‐trifluorobenzene‐carboxamide ( 5h ) was the most active compound, with an IC₅₀ value of 2.84 μm . Compounds 4i , 5h and 5i had lower IC₅₀ values than that found for the clinically employed anticancer drug cisplatin (IC₅₀ = 16.3 μm against MCF‐7). Guanine oxidation studies confirmed that 5h was capable of generating oxidative damage via a reactive oxygen species‐mediated mechanism. Copyright © 2013 John Wiley & Sons, Ltd.     

Nucleophilic addition of bifunctional sulfimidosulfides to platinum(<Emphasis Type="SmallCaps">IV</Emphasis>)-coordinated nitriles

Reactions of the platinum(IV) nitrile complexes [PtCl₄(RCN)₂] (R = Me, CH₂Ph, Ph) with 1,2- and 1,4-PhS(=NH)C₆H₄SPh in CH₂Cl₂ afforded addition products of sulfimides and coordinated nitriles, viz., the [PtCl₄{NH=C(R)N=S(Ph)(C₆H₄SPh)}₂] complexes. The latter were isolated in 75—90% yields and characterized by elemental analysis, positive-ion FAB mass spectrometry, IR spectroscopy, and ¹H and ¹³C¹H NMR spectroscopy. The temperature dependence of the ¹H NMR spectra of the model [PtCl₄{NH=C(R)N=SPh₂}₂] complexes (R = Me, Et) in CD₂Cl₂ studied in a temperature range from +40 to -70 °C demonstrated that E—Z isomerization of the ligands is a dynamic process in a range from +40 to -10 °C. The activation free energy of this process was calculated.Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 8, pp. 1618–1622, August, 2004.     

INVESTIGATION OF THE ATRP OF n-BUTYL METHACRYLATE USING THE Cu（I）／N,N,N＇ N＂ N＂-PENTAMETHYLDIETHYLENETRIAMINE CATALYST SYSTEM

The polymerization of n-butyl methacrylate was investigated using the Atom Transfer Radical Polymerization technique with CuBr and CuCl/N,N,N‘,N“,N“-pentamethyldiethylenetriamine catalytic systems. Various combinations of catalyst systems and initiators were utilized in order to optimize the polymerization conditions and to obtain well-defined polymers (i.e. controlled molecular weights and low polydispersities). It has been found that the optimal initiator for this system is a chlorine-based initiator, when the catalyst used is a Cu(I) salt in conjunction with the N,N,N‘,N“,N“-pentamethyldiethylenetriamine ligand. Bromine-based initiators tend to result in large amounts of initial termination, leading to polymers with less than ideal chain end functionality, even if CuCI is used as the Cu(I) species to invoke the halogen exchange. Additionally, the effects of the polymerization temperature, copper(I) species and the initiator structure were determined.     

Synthesis of porritoxin

A total synthesis following the sequence in Scheme 1 confirms that porritoxin possesses revised structure 3, not the originally assigned 1. A key reaction was the use of iron pentacarbonyl to formylate an aryllithium when DMF and methyl formate proved insufficiently reactive.