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Fraaije MW Kamerbeek NM Heidekamp AJ Fortin R Janssen DB 《The Journal of biological chemistry》2004,279(5):3354-3360
EtaA is a newly identified FAD-containing monooxygenase that is responsible for activation of several thioamide prodrugs in Mycobacterium tuberculosis. It was found that purified EtaA displays a remarkably low activity with the antitubercular prodrug ethionamide. Hinted by the presence of a Baeyer-Villiger monooxygenase sequence motif in the EtaA sequence, we have been able to identify a large number of novel EtaA substrates. It was discovered that the enzyme converts a wide range of ketones to the corresponding esters or lactones via a Baeyer-Villiger reaction, indicating that EtaA represents a Baeyer-Villiger monooxygenase. With the exception of aromatic ketones (phenylacetone and benzylacetone), long-chain ketones (e.g. 2-hexanone and 2-dodecanone) also are converted. EtaA is also able to catalyze enantioselective sulfoxidation of methyl-p-tolylsulfide. Conversion of all of the identified substrates is relatively slow with typical k(cat) values of around 0.02 s(-1). The best substrate identified so far is phenylacetone (K(m) = 61 microM, k(cat) = 0.017 s(-1)). Redox monitoring of the flavin cofactor during turnover of phenylacetone indicates that a step in the reductive half-reaction is limiting the rate of catalysis. Intriguingly, EtaA activity could be increased by one order of magnitude by adding bovine serum albumin. This reactivity and substrate acceptance-profiling study provides valuable information concerning this newly identified prodrug activator from M. tuberculosis. 相似文献
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Annelies Bronckaers Petra Hilkens Yanick Fanton Tom Struys Pascal Gervois Constantinus Politis Wendy Martens Ivo Lambrichts 《PloS one》2013,8(8)
Angiogenesis, the formation of capillaries from pre-existing blood vessels, is a key process in tissue engineering. If blood supply cannot be established rapidly, there is insufficient oxygen and nutrient transport and necrosis of the implanted tissue will occur. Recent studies indicate that the human dental pulp contains precursor cells, named dental pulp stem cells (hDPSC) that show self-renewal and multilineage differentiation capacity. Since these cells can be easily isolated, cultured and cryopreserved, they represent an attractive stem cell source for tissue engineering. Until now, only little is known about the angiogenic abilities and mechanisms of the hDPSC. In this study, the angiogenic profile of both cell lysates and conditioned medium of hDPSC was determined by means of an antibody array. Numerous pro-and anti-angiogenic factors such as vascular endothelial growth factor (VEGF), monocyte chemotactic protein-1 (MCP-1), plasminogen activator inhibitor-1 (PAI-1) and endostatin were found both at the mRNA and protein level. hDPSC had no influence on the proliferation of the human microvascular endothelial cells (HMEC-1), but were able to significantly induce HMEC-1 migration in vitro. Addition of the PI3K-inhibitor and the MEK-inhibitor U0126 to the HMEC-1 inhibited this effect, suggesting that both Akt and ERK pathways are involved in hDPSC-mediated HMEC-1 migration. Antibodies against VEGF also abolished the chemotactic actions of hDPSC. Furthermore, in the chicken chorioallantoic membrane (CAM) assay, hDPSC were able to significantly induce blood vessel formation. In conclusion, hDPSC have the ability to induce angiogenesis, meaning that this stem cell population has a great clinical potential, not only for tissue engineering but also for the treatment of chronic wounds, stroke and myocardial infarctions. LY294002相似文献
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Candy Kumps Annelies Fieuw Pieter Mestdagh Bj?rn Menten Steve Lefever Filip Pattyn Sara De Brouwer Tom Sante Johannes Hubertus Schulte Alexander Schramm Nadine Van Roy Tom Van Maerken Rosa Noguera Valérie Combaret Christine Devalck Frank Westermann Geneviève Laureys Angelika Eggert Jo Vandesompele Katleen De Preter Frank Speleman 《PloS one》2013,8(1)
Neuroblastoma is an embryonic tumor arising from immature sympathetic nervous system cells. Recurrent genomic alterations include MYCN and ALK amplification as well as recurrent patterns of gains and losses of whole or large partial chromosome segments. A recent whole genome sequencing effort yielded no frequently recurring mutations in genes other than those affecting ALK. However, the study further stresses the importance of DNA copy number alterations in this disease, in particular for genes implicated in neuritogenesis. Here we provide additional evidence for the importance of focal DNA copy number gains and losses, which are predominantly observed in MYCN amplified tumors. A focal 5 kb gain encompassing the MYCN regulated miR-17∼92 cluster as sole gene was detected in a neuroblastoma cell line and further analyses of the array CGH data set demonstrated enrichment for other MYCN target genes in focal gains and amplifications. Next we applied an integrated genomics analysis to prioritize MYCN down regulated genes mediated by MYCN driven miRNAs within regions of focal heterozygous or homozygous deletion. We identified RGS5, a negative regulator of G-protein signaling implicated in vascular normalization, invasion and metastasis, targeted by a focal homozygous deletion, as a new MYCN target gene, down regulated through MYCN activated miRNAs. In addition, we expand the miR-17∼92 regulatory network controlling TGFß signaling in neuroblastoma with the ring finger protein 11 encoding gene RNF11, which was previously shown to be targeted by the miR-17∼92 member miR-19b. Taken together, our data indicate that focal DNA copy number imbalances in neuroblastoma (1) target genes that are implicated in MYCN signaling, possibly selected to reinforce MYCN oncogene addiction and (2) serve as a resource for identifying new molecular targets for treatment. 相似文献
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Aims
Because the water status of grapevines strongly affects the quality of the grapes and resulting wine, automated and early drought stress detection is important. Plant measurements are very promising for detecting drought stress, but strongly depend on microclimatic changes. Therefore, conventional stress detection methods require threshold values which define when plants start sensing drought stress. There is however no unique method to define these values. In this study, we propose two techniques that overcome this limitation.Methods
Two statistical methods were used to automatically distinguish between drought and microclimate effects, based on a short preceding full-irrigated period to extract plant behaviour under normal conditions: Unfold Principal Component Analysis (UPCA) and Functional Unfold Principal Component Analysis (FUPCA). Both techniques aimed at detecting when measured sap flow rate or stem diameter variations in grapevine deviated from their normal behaviour due to drought stress.Results
The models based on sap flow rate had some difficulties to detect stress on days with low atmospheric demands, while those based on stem diameter variations did not show this limitation, but ceased detecting stress when the stem diameter levelled off after a period of severe shrinkage. Nevertheless, stress was successfully detected with both approaches days before visible symptoms appeared.Conclusions
UPCA and FUPCA based on plant indicators are therefore very promising for early stress detection. 相似文献8.
Annelies De Paepe Sylvie De Buck Jonah Nolf Els Van Lerberge Ann Depicker 《The Plant journal : for cell and molecular biology》2013,75(1):172-184
Random T–DNA integration into the plant host genome can be problematic for a variety of reasons, including potentially variable transgene expression as a result of different integration positions and multiple T–DNA copies, the risk of mutating the host genome and the difficulty of stacking well‐defined traits. Therefore, recombination systems have been proposed to integrate the T–DNA at a pre‐selected site in the host genome. Here, we demonstrate the capacity of the ?C31 integrase (INT) for efficient targeted T–DNA integration. Moreover, we show that the iterative site‐specific integration system (ISSI), which combines the activities of the CRE recombinase and INT, enables the targeting of genes to a pre‐selected site with the concomitant removal of the resident selectable marker. To begin, plants expressing both the CRE and INT recombinase and containing the target attP site were constructed. These plants were supertransformed with a T–DNA vector harboring the loxP site, the attB sites, a selectable marker and an expression cassette encoding a reporter protein. Three out of the 35 transformants obtained (9%) showed transgenerational site‐specific integration (SSI) of this T–DNA and removal of the resident selectable marker, as demonstrated by PCR, Southern blot and segregation analysis. In conclusion, our results show the applicability of the ISSI system for precise and targeted Agrobacterium‐mediated integration, allowing the serial integration of transgenic DNA sequences in plants. 相似文献
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Michiel?B?HaesekerEmail author Evelien?Pijpers Nicole?HTM?Dukers-Muijrers Patty?Nelemans Christian?JPA?Hoebe Cathrien?A?Bruggeman Annelies?Verbon Valère?J?Goossens 《Immunity & ageing : I & A》2013,10(1):30
Background
Studies about associations of infections with herpes viruses and other pathogens, such as Chlamydia pneumoniae (CP) and Helicobacter pylori (HP) with cardiovascular disease (CVD), diabetes mellitus (DM), frailty and/or mortality are conflicting. Since high levels of antibodies against these pathogens occur in the elderly, the role of these pathogens in morbidity and mortality of vulnerable elderly was explored.Results
Blood samples of 295 community dwelling psycho-geriatric patients were tested for IgG antibodies to herpes simplex virus type 1 and 2, varicella zoster virus, Epstein Barr virus (EBV), cytomegalovirus (CMV), human herpes virus type 6 (HHV6), CP and HP. Frailty was defined with an easy-to-use previously described frailty risk score. Relative risks (RR) with 95% confidence intervals were calculated to evaluate associations between CVD, DM, frailty and pathogens. Pathogens as a predictor for subsequent mortality were tested using Kaplan Meier analyses and Cox proportional hazard models. The mean age was 78 (SD: 6.7) years, 20% died, 44% were defined as frail, 20% had DM and 49% had CVD. Presence of CMV antibody titers was associated with frailty, as shown by using both qualitative and quantitative tests, RR ratio 1.4 (95% CI: 1.003-2.16) and RR ratio 1.5 (95% CI: 1.06-2.30), respectively. High IgG antibody titers of HHV6 and EBV were associated with DM, RR ratio 3.3 (95% CI: 1.57-6.49). None of the single or combined pathogens were significantly associated with mortality and/or CVD.Conclusions
Prior CMV infection is associated with frailty, which could be in line with the concept that CMV might have an important role in immunosenescence, while high IgG titers of HHV6 and EBV are associated with DM. No association between a high pathogen burden and morbidity and/or mortality could be demonstrated.10.
Henk Schierbeek Annelies H.C. Goris Koen F.M. Joosten Imke van Kessel Eva Corpeleijn Pieter J.J. Sauer 《Obesity (Silver Spring, Md.)》2013,21(9):1877-1883
Objectives: To assess validity evidence of TracmorD to determine energy used for physical activity in 3‐4‐year‐old children. Design and Methods: Participants were randomly selected from GECKO Drenthe cohort (n = 30, age 3.4 ± 0.3 years). Total energy expenditure (TEE) was measured using the doubly labeled water method. Sleeping metabolic rate (SMR) was measured by indirect calorimetry (Deltatrac). TEE and SMR were used to calculate physical activity level (PAL) and activity energy expenditure (AEE). Physical activity was monitored using a DirectLife triaxial accelerometer, TracmorD with activity counts per minute (ACM) and activity counts per day (ACD) as outcome measures. Results: The best predictor for PAL was ACM with gender and weight, the best predictor for AEE was ACM alone (backward regression, R2 = 0.50, P = 0.010 and R2 = 0.31, P = 0.011, respectively). With ACD, the prediction model for PAL included ACD, height, gender, and sleep duration (R2 = 0.48, P = 0.033), the prediction model for AEE included ACD, gender and sleep duration (R2 = 0.39, P = 0.042). The accelerometer was worn for 5 days, but 3 days did not give a different estimated PAL. Conclusion: TracmorD provides moderate‐to‐strong validity evidence that supports its use to evaluate energy used for physical activity in 3‐4‐year‐old children. 相似文献