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A novel scheme for precise diagnostics and effective stabilization of currents in a fuel cell stack
(2010)
Lightning safety guidelines
(2010)
Realization of a calorimetric gas sensor on polyimide foil for applications in aseptic food industry
(2010)
Normative Regulations
(2010)
Hybrid control for autonomous systems — Integrating learning, deliberation and reactive control
(2010)
C-terminal truncation of a metagenome-derived detergent protease for effective expression in E. coli
(2010)
Recently, a new alkaline protease named HP70 showing highest homology to extracellular serine proteases of Stenotrophomonas maltophilia and Xanthomonas campestris was found in the course of a metagenome screening for detergent proteases (Niehaus et al., submitted for publication). Attempts to efficiently express the enzyme in common expression hosts had failed. This study reports on the realization of overexpression in Escherichia coli after structural modification of HP70. Modelling of HP70 resulted in a two-domain structure, comprising the catalytic domain and a C-terminal domain which includes about 100 amino acids. On the basis of the modelled structure the enzyme was truncated by deletion of most of the C-terminal domain yielding HP70-C477.
This structural modification allowed effective expression of active enzyme using E. coli BL21-Gold as the host. Specific activity of HP70-C477 determined with suc-l-Ala-l-Ala-l-Pro-l-Phe-p-nitroanilide as the substrate was 30 ± 5 U/mg compared to 8 ± 1 U/mg of the native enzyme. HP70-C477 was most active at 40 °C and pH 7–11; these conditions are prerequisite for a potential application as detergent enzyme. Determination of kinetic parameters at 40 °C and pH = 9.5 resulted in KM = 0.23 ± 0.01 mM and kcat = 167.5 ± 3.6 s⁻¹. MS-analysis of peptide fragments obtained from incubation of HP70 and HP70-C477 with insulin B indicated that the C-terminal domain influences the cleavage preferences of the enzyme. Washing experiments confirmed the high potential of HP70-C477 as detergent protease.
Epilepsy
(2010)
Objective: As high-field cardiac MRI (CMR) becomes more widespread the propensity of ECG to interference from electromagnetic fields (EMF) and to magneto-hydrodynamic (MHD) effects increases and with it the motivation for a CMR triggering alternative. This study explores the suitability of acoustic cardiac triggering (ACT) for left ventricular (LV) function assessment in healthy subjects (n=14). Methods: Quantitative analysis of 2D CINE steady-state free precession (SSFP) images was conducted to compare ACT’s performance with vector ECG (VCG). Endocardial border sharpness (EBS) was examined paralleled by quantitative LV function assessment. Results: Unlike VCG, ACT provided signal traces free of interference from EMF or MHD effects. In the case of correct Rwave recognition, VCG-triggered 2D CINE SSFP was immune to cardiac motion effects—even at 3.0 T. However, VCG-triggered 2D SSFP CINE imaging was prone to cardiac motion and EBS degradation if R-wave misregistration occurred. ACT-triggered acquisitions yielded LV parameters (end-diastolic volume (EDV), endsystolic volume (ESV), stroke volume (SV), ejection fraction (EF) and left ventricular mass (LVM)) comparable with those derived fromVCG-triggered acquisitions (1.5 T: ESVVCG=(56± 17) ml, EDVVCG=(151±32)ml, LVMVCG=(97±27) g, SVVCG=(94± 19)ml, EFVCG=(63±5)% cf. ESVACT= (56±18) ml, EDVACT=(147±36) ml, LVMACT=(102±29) g, SVACT=(91± 22) ml, EFACT=(62±6)%; 3.0 T: ESVVCG=(55±21) ml, EDVVCG=(151±32) ml, LVMVCG=(101±27) g, SVVCG=(96±15) ml, EFVCG=(65±7)% cf. ESVACT=(54±20) ml, EDVACT=(146±35) ml, LVMACT= (101±30) g, SVACT=(92±17) ml, EFACT=(64±6)%). Conclusions: ACT’s intrinsic insensitivity to interference from electromagnetic fields renders
Mouse nongenotoxic hepatocarcinogens phenobarbital (PB) and chlordane induce hepatomegaly characterized by hypertrophy and hyperplasia. Increased cell proliferation is implicated in the mechanism of tumor induction. The relevance of these tumors to human health is unclear. The xenoreceptors, constitutive androstane receptors (CARs), and pregnane X receptor (PXR) play key roles in these processes. Novel “humanized” and knockout models for both receptors were developed to investigate potential species differences in hepatomegaly. The effects of PB (80 mg/kg/4 days) and chlordane (10 mg/kg/4 days) were investigated in double humanized PXR and CAR (huPXR/huCAR), double knockout PXR and CAR (PXRKO/CARKO), and wild-type (WT) C57BL/6J mice. In WT mice, both compounds caused increased liver weight, hepatocellular hypertrophy, and cell proliferation. Both compounds caused alterations to a number of cell cycle genes consistent with induction of cell proliferation in WT mice. However, these gene expression changes did not occur in PXRKO/CARKO or huPXR/huCAR mice. Liver hypertrophy without hyperplasia was demonstrated in the huPXR/huCAR animals in response to both compounds. Induction of the CAR and PXR target genes, Cyp2b10 and Cyp3a11, was observed in both WT and huPXR/huCAR mouse lines following treatment with PB or chlordane. In the PXRKO/CARKO mice, neither liver growth nor induction of Cyp2b10 and Cyp3a11 was seen following PB or chlordane treatment, indicating that these effects are CAR/PXR dependent. These data suggest that the human receptors are able to support the chemically induced hypertrophic responses but not the hyperplastic (cell proliferation) responses. At this time, we cannot be certain that hCAR and hPXR when expressed in the mouse can function exactly as the genes do when they are expressed in human cells. However, all parameters investigated to date suggest that much of their functionality is maintained.