Enzymatic Addition of Alcohols to Terpenes by Squalene Hopene Cyclase Variants

Squalene–hopene cyclases (SHCs) catalyze the polycyclization of squalene into a mixture of hopene and hopanol. Recently, amino-acid residues lining the catalytic cavity of the SHC from Alicyclobacillus acidocaldarius were replaced by small and large hydrophobic amino acids. The alteration of leucine 607 to phenylalanine resulted in increased enzymatic activity towards the formation of an intermolecular farnesyl–farnesyl ether product from farnesol. Furthermore, the addition of small-chain alcohols acting as nucleophiles led to the formation of non-natural ether-linked terpenoids and, thus, to significant alteration of the product pattern relative to that obtained with the wild type. It is proposed that the mutation of leucine at position 607 may facilitate premature quenching of the intermediate by small alcohol nucleophiles. This mutagenesis-based study opens the field for further intermolecular bond-forming reactions and the generation of non-natural products.     

Optimierung der Habitatmodellierung an gro?en Flie?gew?ssern unter Berücksichtigung von flussmorphologischen Prozessen auf unterschiedlichen Skalen

In light of the various anthropogenic pressures on our river systems (flood protection, navigation, recreational fisheries) and current legal requirements (European Water Framework Directive), the need for integrative evaluation tools has become increasingly apparent. The so-called habitat modeling approach addresses these integrative efforts to quantify the changing environment for target species (fish, macro-invertebrates), taking into account hydrological and/or morphological changes. Over the course of extensive monitoring on the Danube east of Vienna this specific evaluation tool has been adapted and improved to allow the assessment of river engineering measures in future, especially before their implementation and to determine their potential impacts on the ecological conditions.     

Crystal Structure Determination and Mutagenesis Analysis of the Ene Reductase NCR

The crystal structure of the “ene” nicotinamide‐dependent cyclohexenone reductase (NCR) from Zymomonas mobilis (PDB ID: 4A3U) has been determined in complex with acetate ion, FMN, and nicotinamide, to a resolution of 1.95 Å. To study the activity and enantioselectivity of this enzyme in the bioreduction of activated α,β‐unsaturated alkenes, the rational design methods site‐ and loop‐directed mutagenesis were applied. Based on a multiple sequence alignment of various members of the Old Yellow Enzyme family, eight single‐residue variants were generated and investigated in asymmetric bioreduction. Furthermore, a structural alignment of various ene reductases predicted four surface loop regions that are located near the entrance of the active site. Four NCR loop variants, derived from loop‐swapping experiments with OYE1 from Saccharomyces pastorianus, were analysed for bioreduction. The three enzyme variants, P245Q, D337Y and F314Y, displayed increased activity compared to wild‐type NCR towards the set of substrates tested. The active‐site mutation Y177A demonstrated a clear influence on the enantioselectivity. The loop‐swapping variants retained reduction efficiency, but demonstrated decreased enzyme activity compared with the wild‐type NCR ene reductase enzyme.     

Low prevalence of coronary artery spasm in patients with normal coronary angiograms and unexplained ventricular fibrillation

AIMS: The aetiology of ventricular fibrillation in patients without identifiable structural heart disease is unknown. Recently, high prevalence of silent ischaemia due to coronary artery spasm has been reported in such patients. However, in at least one report, all patients had non-critical coronary artery lesions. Identification of coronary artery spasm as the underlying aetiology of ventricular fibrillation has important therapeutic implications. METHODS AND RESULTS: We performed ergonovine provocation tests in 18 patients (14 males, and four females; mean age, 36 years) with documented ventricular fibrillation in the absence of identifiable structural heart disease who had undergone aborted sudden death. In group I (n = 7) ergonovine provocation tests were performed at a mean interval of 31 months (range 21-42 months) after the index episode. These patients had already received an implantable cardioverter defibrillator, after failed electrophysiologically guided antiarrhythmic therapy. In group II (n = 11) the ergonovine provocation test was performed prospectively as part of the diagnostic evaluation. All patients were off antiarrhythmic drugs, calcium entry or beta-adrenoceptor blockers at the time of the ergonovine provocation test. Ergonovine was administered intravenously as a bolus injection, beginning with 0.05 mg followed every 3 min by incremental doses up to a cumulative maximum dose of 0.45 mg. Predefined end-points were (1) recording of ischaemic ST segment shifts of > or = 1 mm in at least two corresponding leads of the 12-lead electrocardiogram; (2) induction of ventricular tachycardia or ventricular fibrillation; and (3) administration of a cumulative dose of 0.45 mg. A positive response to ergonovine was seen in only one patient (5%) in group I in whom there developed ST segment elevation without angina and a short burst of rapid ventricular tachycardia. CONCLUSIONS: This study found a low prevalence of coronary artery spasm in patients with aborted sudden death resulting from documented ventricular fibrillation and non-apparent underlying heart disease. All patients had normal coronary angiograms and a negative history for spontaneous episodes of chest pain. The mechanism of arrhythmogenesis in such patients remains largely unknown.     

A 1.8-V 91-dB DR second-order ΣΔ modulator in 0.18-μm CMOS technology

This paper deals with the implementation of a second-order ΣΔ modulator in 0.18-μm CMOS technology. The analog-to-digital converter structure combines a 1-bit approach along with a high oversampling ratio (OSR). A silicon circuit prototype, including the modulator itself, a current reference, and the clock signals generator, was designed to operate with a 1.8-V supply, fabricated and tested. Measured values of 87 dB and 91 dB were obtained for the signal-to-noise-plus-distortion ratio (SNDR) and the dynamic range (DR), respectively, for a clock frequency of 8 MHz and an OSR of 256. The effective number of bits (ENOB) was above 14. The experimental performance of the ΣΔ modulator maintains a good level over a modulator clock range higher than 16 MHz, featuring an ENOB equal to 13 at 16 MHz.     

Prediction in road safety studies: An empirical inquiry

Studies about the road safety effect of interventions are usually retrospective quasiexperiments. In these, one key task is to predict what would have been the safety of the treated group without the intervention. Such predictions can be made by several methods, one of which is to use a “comparison group.” We use 26 yearly counts of reported injury accidents for the Canadian provinces to examine which of several simple methods of prediction would have historically predicted best. We find that the use of more data does not always improve prediction. How well one predicts depends not only on the amount of data used but also on the extent to which the prediction method is in accord with the unknown time trend behind the accident counts. We also find that the use of a comparison group to predict is not always better than predicting that this year's count will be the same as last year's. In addition, the intuitive notion that a good comparison group is that which is thought similar to the treated group is too simple. Both similarity and size (as measured by the number of accidents) are important. Moreover, whatever preconceived notions of similarity we had, were contradicted by the data. If the history of accident counts on the treatment group and on several possible comparison groups is available, a simple method to select the most suitable comparison group is suggested.     

Optimierung von Wasserkraft und Ökologie bei Erneuerung oder Revitalisierung bestehender Anlagen – „win-win“-Ansatz

In Austria, using hydropower has a centuries-long tradition. Whereas we first placed mills along our rivers to put water’s power to use, today we have over 5,200 water power stations of various types, from the smallest facilities to pumped-storage hydroelectric plants in the Alps. Today, water power accounts for roughly 60 % of our domestic power production, though the various types of power stations used have diverse ecological effects. This article presents a “win-win” approach to improving both hydropower production and ecological outcomes on the basis of the Kamp River in Lower Austria. The Kamp is not only home to a pumped-storage hydroelectric plant and the major plants Ottenstein, Dobra-Krumberg and Thurnberg-Wegscheid, but also to 14 smaller plants with turbines that are between 25 and 75 years old, many of them with energy conversion efficiency ratios of well under 0.7. Using the case study of a small water power plant, the chances of achieving a win-win situation are presented in detail. In this context, replacing the turbines is a major aspect of improving these plants. Extensive analyses have demonstrated that balanced planning approaches can yield both higher efficiency and better conditions for aquatic ecology, thanks to increased residual flows. Further, energy calculations and numerical habitat simulations have confirmed that a win-win situation is feasible for the small power plant studied, especially if Kaplan turbines are used. For example, they show that, despite the increased residual flow of up to 800 l/s, a considerable performance increase over the status quo is possible (maximum performance increase of 17 % without considering residual flow). If the residual flow was only 400 l/s, which tests indicate would mean an improvement in the habitat conditions for all ages of the dominant fish species (the brown trout) of > 50 %, a performance increase of 8 % would be the result. Thanks to the generally applicable methodology used, the win-win approach presented here can broadly be used in the optimization, efficiency improvement and revitalization of current hydropower plants, producing both more electricity and higher residual flow levels to improve local ecological conditions. In order to make this win-win situation a reality, innovative and integrative financial models for e.g. new turbines related to the fields of green energy production and environmental protection will be essential.     

Crash reductions related to traffic signal removal in Philadelphia

The effect on intersection crashes of converting one-way street intersections in Philadelphia from signal to multiway stop sign control was estimated. Using crash and traffic volume data for a comparison group, regression models were computed to represent the normal crash experience of signal controlled intersections of one-way streets, by impact type, as a function of traffic volume. An empirical Bayesian procedure was used to estimate what would have been the expected number of crashes at the converted intersections had they not been converted. The empirical Bayesian estimates were compared with actual counts of crashes after conversion. Estimates were obtained for different classes of crashes categorized by impact type, day/night condition, and impact severity. Aggregate results indicate that replacing signals by multiway stop signs on one-way streets is associated with a reduction in crashes of approximately 24%, combining all severities, light conditions, and impact types.