Development of a whole-cell biocatalyst co-expressing P450 monooxygenase and glucose dehydrogenase for synthesis of epoxyhexane

Oxygenases-based Escherichia coli whole-cell biocatalyst can be applied for catalysis of various commercially interesting reactions that are difficult to achieve with traditional chemical catalysts. However, substrates and products of interest are often toxic to E. coli, causing a disruption of cell membrane. Therefore, organic solvent-tolerant bacteria became an important tool for heterologous expression of such oxygenases. In this study, the organic solvent-tolerant Bacillus subtilis 3C5N was developed as a whole-cell biocatalyst for epoxidation of a toxic terminal alkene, 1-hexene. Comparing to other hosts tested, high level of tolerance towards 1-hexene and a moderately hydrophobic cell surface of B. subtilis 3C5N were suggested to contribute to its higher 1,2-epoxyhexane production. A systematic optimization of reaction conditions such as biocatalyst and substrate concentration resulted in a 3.3-fold increase in the specific rate. Co-expression of glucose dehydrogenase could partly restored NADPH-regenerating ability of the biocatalyst (up to 38?% of the wild type), resulting in approximately 53?% increase in specific rate representing approximately 22-fold increase in product concentration comparing to that obtained prior to an optimization.     

Concentration of live pico- and nanoplankton by means of tangential flow filtration

A tangential flow filtration system for concentrating live pico-and nanoplankton was tested on natural seawater (<20 µm)from Kiel Bight. Twenty- to thirty-fold concentration of samplesranging from 79 to 152 dm³ was easily achieved, the time necessarydepending on sample volume, pumping pressure and filtrate/retentateratio. Losses of different kinds of cells in relation to theconcentration factor were quantified. Recovery rates were negativelycorrelated with the frequency of sample recirculation throughthe filtration unit.     

Why do organisms produce gametes of only two different sizes? Some theoretical aspects of the evolution of anisogamy

This paper extends the population genetic model of (the evolution of) anisogamy of Charlesworth (1978), which is based on the model of Parker, Baker & Smith (1972). The effect of parthenogenesis on the evolution of anisogamy is examined; this effect turns out to be only quantitative. Furthermore, the problem of the occurrence of only two different gamete sizes is considered. It is shown that a stable polymorphism with three different gamete sizes cannot exist. This result is robust to changes in the mating structure (random or disassortative gamete fusion) and to changes in the mode of reproduction (only sexual or partially parthenogenetic).     

Bioanalysis of aplidine, a new marine antitumoral depsipeptide, in plasma by high-performance liquid chromatography after derivatization with trans-4′-hydrazino-2-stilbazole

A sensitive bio-analytical assay in plasma of the depsipeptide aplidine is reported, based on reversed-phase liquid chromatography and fluorescence detection of the trans-4′-hydrazino-2-stilbazole (4′H2S) derivative of the analyte. At ambient temperature, two conformations of the depsipeptide are observed in solution due to cis–trans isomerism at the proline–pyruvoyl peptide bond. Aplidine is isolated from the matrix by solid-phase extraction on an octadecyl modified silica stationary phase. After evaporation of the acetone eluate, a derivatization with 4′H2S is performed in a water–acetonitrile mixture at pH 4. The reaction mixture is injected directly into the chromatograph and the analyte is quantified by fluorescence detection at 410 and 560 nm for excitation and emission, respectively. The method has been validated in the 2–100 ng/ml-range, 2 ng/ml being the lower limit of quantification. Precision and accuracy both meet the current requirements for a bioanalytical assay. The identity of the 4′H2S reaction products of aplidine have been confirmed by mass spectrometric analysis. Finally, the method has been employed for a pilot pharmacokinetic study of aplidine in mice which demonstrated its usefulness for pharmacological research.     

Structure of the human type I DNA topoisomerase gene

We describe the molecular organization of the human gene coding for type I DNA topoisomerase. The coding sequence is split into 21 exons distributed over at least 85 kilobase pairs (kb) of human genomic DNA. The sizes of the 20 introns vary widely between 0.2 and at least 30 kb and all contain the sequence elements known to be required for pre-mRNA splicing. Several of the intron sequences separate exons encoding parts of the enzyme that are highly conserved between human and yeast suggesting that at least some of the exons may code for individual, structurally, or functionally important domains of the enzyme. We also describe the promoter sequence of the human topoisomerase I gene and show that it is composed of distinct functional elements.     

COMPOSITION OF VOLATILES IN RELATION TO TAXONOMY OF AMERICAN ALLIUMS

The North American species of Allium exclusive of A. schoenoprasum and A. tricoccum of Old World affinity are grouped on the basis of morphological similarity into eight discontinuous species alliances typified by A. acuminatum, A. campanulatum, A. canadense, A. cernuum, A. falcifolium, A. kunthii, A. sanbornii, and A. validum, respectively. Representatives of each of these alliances were compared with respect to volatile constituents responsible for characteristic odors by means of gas chromatography. Results indicate that these volatiles provide evidence of relationship useful in the classification of alliums. Uniformity was found in composition of volatiles in the representatives of the A. canadense, A. cernuum, A. kunthii, and A. sanbornii alliances. Variation was observed in the A. acuminatum, A. campanulatum, and A. falcifolium alliances. A. validum was the only species of its alliance studied. Vapors of A. validum contain mostly n-propyl sulfides (onion-like odor) as does the cultivated A. cepa. Methyl sulfides (cabbage-like odor) predominate in the A. sanbornii alliance. A few species of the A. acuminatum and A. falcifolium alliances contain mainly allyl sulfides (garlic-like odor).