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Saale, Germany

Behr S.,Ludwig Maximilians University of Munich | Fried L.,Ludwig Maximilians University of Munich | Fried L.,BioSolutions GmbH | Jung K.,Ludwig Maximilians University of Munich
Journal of Bacteriology | Year: 2014

When carbon sources become limiting for growth, bacteriamust choose which of the remaining nutrients should be used first. We have identified a nutrient-sensing signaling network in Escherichia coli that is activated at the transition to stationary phase. The network is composed of the two histidine kinase/response regulator systems YehU/YehT and YpdA/YpdB and their target proteins, YjiY and YhjX (both of which aremembrane-integrated transporters). The peptide/amino acid-responsive YehU/YehT systemwas found to have a negative effect on expression of the target gene, yhjX, of the pyruvate-responsive YpdA/YpdB system, while the YpdA/YpdB systemstimulated expression of yjiY, the target of the YehU/YehT system. These effects were confirmed in mutants lacking any of the genes for the three primary components of either system. Furthermore, an in vivo interaction assay based on bacterial adenylate cyclase detected heteromeric interactions between themembrane-bound components of the two systems, specifically, between the two histidine kinases and the two transporters, which is compatible with the formation of a larger signaling unit. Finally, the carbon storage regulator A (CsrA) was shown to be involved in posttranscriptional regulation of both yjiY and yhjX. © 2014, American Society for Microbiology. Source


Lassak J.,Ludwig Maximilians University of Munich | Fried L.,Ludwig Maximilians University of Munich | Fried L.,BioSolutions GmbH | Jung K.,Ludwig Maximilians University of Munich
BioSpektrum | Year: 2014

ß-galactosidase encoded by lacZ remains a popular reporter enzyme. Here, we present a convenient tool box to facilitate rapid construction of reporter lacZ fusions. The first tool enables the simple generation of chromosomally encoded reporters within the Escherichia coli lac operon using Red®/ET® recombination. The second tool allows the introduction of lacZ reporter fusions into the chromosome by a single-crossover method. The third tool comprises four pBBR1-based broad-host-range vectors to derive transcriptional and translational lacZ fusions. © 2014, Springer-Verlag Berlin Heidelberg. Source


Kommera H.,Martin Luther University of Halle Wittenberg | Kaluderovic G.N.,Martin Luther University of Halle Wittenberg | Dittrich S.,Martin Luther University of Halle Wittenberg | Kalbitz J.,BioSolutions GmbH | And 3 more authors.
Bioorganic and Medicinal Chemistry Letters | Year: 2010

Synthesis and antiproliferative activity of eight new derivatives of betulinic acid (1) and betulin (2) are described. The compounds were tested against fifteen tumor cell lines. The toxicity against normal human fibroblasts and the mode of cell death on lung cancer cell line induced by the most active compounds 9 (bis(ethylcarbamate)betulin) and 11 (3-O-ethylcarbamate of 28-O-acetylbetulin) was investigated. Caspase 3 activity on lung cancer cell line (A549) was determined for 1, 5 (3-O-ethylcarbamate of betulinic acid), 9 and 11. All derivatives exerted a dose dependent antiproliferative action at micromolar concentrations toward target tumor cell lines. Treatment of lung cancer cells for 24 h with 9 and 11 induced apoptosis, as observed by the appearance of a typical ladder pattern in the DNA fragmentation assay. © 2010 Elsevier Ltd. All rights reserved. Source


Kommera H.,Martin Luther University of Halle Wittenberg | Kaluderovic G.N.,Martin Luther University of Halle Wittenberg | Kaluderovic G.N.,University of Belgrade | Kalbitz J.,BioSolutions GmbH | And 2 more authors.
European Journal of Medicinal Chemistry | Year: 2010

In the present investigations five new derivatives of betulinic and betulonic acid were synthesized and the effect of this structural variations on anticancer activity was studied and discussed. The anti-proliferative activity of betulinic and betulonic acid derivatives was studied against eight tumor cell lines of different histogenic origin. The derivatives exerted a dose dependent antiproliferative action at micromolar concentrations toward target tumor cell lines. The apoptotic mode of cell death on colon cancer cell line HT-29 was induced by the most active compounds 5,2-amino-3-hydroxy-2-(hydroxy-methyl) propyl (3-O-acetyl)betulinate,and 9,2-amino-3-hydroxy-2-(hydroxymethyl)propyl betulonate. Treatment of HT-29 cells with 5 and 9 induced apoptosis,as observed by dye exclusion test (trypan blue) and by the appearance of a typical ladder pattern in the DNA fragmentation assay and FITC annexin V assay. Cell cycle perturbations caused by compound 5 are also presented. © 2010 Elsevier Masson SAS. All rights reserved. Source


Koch B.,Probiodrug | Koch B.,BioSolutions GmbH | Kolenko P.,Martin Luther University of Halle Wittenberg | Buchholz M.,Probiodrug | And 9 more authors.
Biochemistry | Year: 2012

Glutaminyl cyclases (QCs), which catalyze the formation of pyroglutamic acid (pGlu) at the N-terminus of a variety of peptides and proteins, have attracted particular attention for their potential role in Alzheimer's disease. In a transgenic Drosophila melanogaster (Dm) fruit fly model, oral application of the potent competitive QC inhibitor PBD150 was shown to reduce the burden of pGlu-modified Aβ. In contrast to mammals such as humans and rodents, there are at least three DmQC species, one of which (isoDromeQC) is localized to mitochondria, whereas DromeQC and an isoDromeQC splice variant possess signal peptides for secretion. Here we present the recombinant expression, characterization, and crystal structure determination of mature DromeQC and isoDromeQC, revealing an overall fold similar to that of mammalian QCs. In the case of isoDromeQC, the putative extended substrate binding site might be affected by the proximity of the N-terminal residues. PBD150 inhibition of DromeQC is roughly 1 order of magnitude weaker than that of the human and murine QCs. The inhibitor binds to isoDromeQC in a fashion similar to that observed for human QCs, whereas it adopts alternative binding modes in a DromeQC variant lacking the conserved cysteines near the active center and shows a disordered dimethoxyphenyl moiety in wild-type DromeQC, providing an explanation for the lower affinity. Our biophysical and structural data suggest that isoDromeQC and human QC are similar with regard to functional aspects. The two Dm enzymes represent a suitable model for further in-depth analysis of the catalytic mechanism of animal QCs, and isoDromeQC might serve as a model system for the structure-based design of potential AD therapeutics. © 2012 American Chemical Society. Source

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