Highly Active and Diastereoselective N,O‐ and N,N‐Yttrium Complexes for Intramolecular Hydroamination

The intramolecular hydroamination of aminoalkynes and unactivated aminoalkenes catalyzed by yttrium N,O‐ and N,N‐complexes has been investigated. The N,N‐yttrium complexes are highly active, catalyzing the conversion of a wide range of terminal aminoalkenes at room temperature, and internal aminoalkenes at elevated temperature, to yield pyrrolidine and piperidine products in high yields. A high diastereoselectivity of up to 23:1 is observed at 0 °C with 1‐methyl‐4‐pentenylamine as substrate.     

Efficient Trifluoromethylation of Activated and Non‐Activated Alkenyl Halides by Using (Trifluoromethyl)trimethylsilane

An efficient method for the trifluoromethylation of halogenated double bonds by using in situ generated “trifluoromethyl copper” is described. Herein, the most common trifluoromethyl source, (trifluoromethyl)trimethylsilane, was converted selectively into “trifluoromethyl copper” by using copper iodide as copper source and potassium fluoride as promoter. In 1,3‐dimethyl‐3,4,5,6‐tetrahydro‐2(1H)‐pyrimidinone (DMPU) as chelating solvent the trifluoromethylation of activated and non‐activated alkenyl halides occurred mostly in good to excellent yields in up to a gram scale.     

Remarkable findings concerning PBDEs in the terrestrial top-predator red fox (Vulpes vulpes)

In the present study, we have analyzed muscle, liver, and adipose tissue of 33 red foxes from Belgium for their content of polybrominated diphenyl ethers (PBDEs). Median sums of seven tri- to hepta-BDEs (BDE 28, BDE 47, BDE 99, BDE 100, BDE 153, BDE 154, and BDE 183) were 2.2, 2.4, and 3.4 ng/g lipid weight in adipose tissue, liver, and muscle, respectively. These levels were lower than those found in various species of voles and mice, the main prey species of the red fox. This is probably related to the high capacity of the foxes to metabolize and eliminate lower brominated congeners. BDE 209 generally dominated the PBDE congener profiles in the red fox samples. In samples containing BDE 209, this congener contributed, on the average, approximately 70% to the total PBDE content. BDE 209 was measured in concentrations as high as 760 ng/g lipid weight in the liver, but the detection frequency was not more than 40%. In animals with the highest BDE 209 levels, this congener was detected in muscle, liver, as well as in adipose tissue. Other abundant congeners were BDE 153 and BDE 47, which prevail in other terrestrial species. The particular PBDE congener profile observed in the red fox resembles that seen in grizzly bears from Canada, but differs from those previously reported for terrestrial avian species. Our data confirms unambiguously that BDE 209 does bioaccumulate in terrestrial top predators, such as the red fox.     

On-line reaction monitoring in the liquid phase using two mid-infrared quantum cascade lasers simultaneously

On-line monitoring of a model reaction was performed by employing two pulsed mid-infrared Fabry-Pérot quantum cascade lasers (QCL). The emission maxima of the QCLs were located at 1393 and 1080 cm(-1). An optical system of parabolic mirrors and a ZnSe beam splitter combined the two laser beams and allowed a transmission cell to be probed with both QCLs simultaneously. The reaction mixture was pumped continuously through a cell that had an optical path of 48 microm. This dual QCL system allowed fast absorption measurements of the reaction mixture at two distinct wavenumbers. The reaction under study was the oxidation of sulfite to sulfate with hydrogen peroxide acting as oxidant. On-line measurements of the chemical reaction allowed direct, real-time monitoring of sulfate formation and hydrogen peroxide depletion.     

Parallel manipulation of bifunctional DNA molecules on structured surfaces using kinesin-driven microtubules

We have developed a technique to manipulate bifunctional DNA molecules: One end is thiolated to bind to a patterned gold surface and the other end is biotinylated to bind to a microtubule gliding over a kinesin-coated surface. We found that DNA molecules can be stretched and overstretched between the gold pads and the motile microtubules, and that they can form dynamic networks. This serves as a proof-of-principle that biological machineries can be used in vitro to accomplish the parallel formation of structured DNA templates that will have applications in biophysics and nanoelectronics.     

Natural products from marine organisms and their associated microbes

The marine environment is distinguished by unique groups of organisms being the source of a wide array of fascinating structures. The enormous biodiversity of marine habitats is mirrored by the molecular diversity of secondary metabolites found in marine animals, plants and microbes. The recognition that many marine invertebrates contain endo- and epibiotic microorganisms and that some invertebrate-derived natural products are structurally related to bacterial metabolites suggests a microbial origin for some of these compounds. Other marine natural products, however, are clearly located in invertebrate tissue and microbial involvement in the biosynthetic process seems unlikely. The complexity of associations in marine organisms, especially in sponges, bryozoans and tunicates, makes it extremely difficult to definitively state the biosynthetic source of many marine natural products or to deduce their ecological significance. Whereas many symbiotic marine microorganisms cannot be isolated and cultured, numerous epi- and endobiotic marine fungi produce novel secondary metabolites in laboratory cultures. The potent biological activity of many marine natural products is of relevance for their ecological function but is also the basis of their biomedical importance.     

C-terminal modifications of a protein by UAG-encoded incorporation of puromycin during in vitro protein synthesis in the absence of release factor 1

Deactivation of release factor 1 by polyclonal antibodies in an in vitro translation system, which was used to express the esterase gene, led to the reversible elimination of naturally occurring termination. This technique allowed the antibiotic puromycin to be used as an acceptor substrate for the peptidyl residue in the peptidyl-transferase reaction. This resulted in more than 80 % yield of protein with C-terminally incorporated puromycin. pCpPuromycin that was either conjugated with the Cy3 fluorophor or biotin by N4 alkylation of cytosine, also acted as an acceptor substrate for the peptidyl-transferase reaction and was incorporated into the protein C terminus. The resulting conjugates possessed Cy3-specific fluorescence and affinity to streptavidin-coated surfaces, respectively. This left the enzymatic activity of the reporter protein unaffected. It was also shown that extension of puromycin on its 5'-hydroxyl end by up to ten deoxyoligonucleotides also allowed conjugation with the C terminus of in vitro translated protein when RF1-dependent termination was suppressed. However, the conjugation yield decreased upon addition of more than six nucleotides.     

A unique mechanism for methyl ester formation via an amide intermediate found in myxobacteria

Secondary metabolism involves a broad diversity of biochemical reactions that result in a wide variety of biologically active compounds. Terminal amide formation during the biosynthesis of the myxobacterial electron-transport inhibitor, myxothiazol, was analyzed by heterologous expression of the unique nonribosomal-peptide synthetase, MtaG, and incubation with a synthesized substrate mimic. These experiments provide evidence that the terminal amide is formed from a carrier protein-bound myxothiazol acid that is thioesterified to MtaF. This intermediate is transformed to an amide by extension with glycine and subsequent oxidative cleavage by MtaG. The final steps of melithiazol assembly involve a highly similar protein-bound intermediate (attached to MelF, a homologue of MtaF), which is transformed to an amide by MelG (homologue of MtaG). In this study, we also show that the amide moiety of myxothiazol A can be hydrolyzed in vivo to the formerly unknown free myxothiazol acid by heterologous expression of melJ in the myxothiazol producer Stigmatella aurantiaca DW4/3-1. The methyltransferase MelK can finally methylate the acid to give rise to the methyl ester, which is produced as the final product in the melithiazol A biosynthetic pathway. These experiments clarify the role of MelJ and MelK during melithiazol assembly.     

Polyaza crown ethers as non-nucleosidic building blocks in DNA conjugates: synthesis and remarkable stabilization of dsDNA

The synthesis of amphiphilic polyaza crown ether monomers X (benzyl-substituted), Y (palmityl-substituted) and Z (cholesteryl-substituted) and their incorporation into oligonucleotides are described. Their effects on thermal duplex stability were investigated by UV melting curve analysis in different alkaline metal buffer solutions. Thermal-denaturation experiments showed remarkable stabilization of dsDNA by polyaza crown ether monomers when incorporated in opposite positions. The series of polyaza crown ether monomers (X, Y and Z) with different overall lipophilicities showed a trend of increased stability of the corresponding dsDNA with increasing lipophilicity of the polyaza crown ether monomer. Multiple incorporations of benzyl-substituted polyaza crown ether monomer X as dangling ends on both sides of dsDNA resulted in strongly increased stability of the corresponding duplex.