De novo genome assembly and analysis unveil biosynthetic and metabolic potentials of Pseudomonas fragi A13BB.

OBJECTIVES: The role of rhizosphere microbiome in supporting plant growth under biotic stress is well documented. Rhizobacteria ward off phytopathogens through various mechanisms including antibiosis. We sought to recover novel antibiotic-producing bacterial strains from soil samples collected from the rhizosphere. Pseudomonas fragi A13BB was recovered as part of this effort, and the whole genome was sequenced to facilitate mining for potential antibiotic-encoding biosynthetic gene clusters. DATA DESCRIPTION: Here, we report the complete genome sequence of P. fragi A13BB obtained from de novo assembly of Illumina MiSeq and GridION reads. The 4.94 Mb genome consists of a single chromosome with a GC content of 59.40%. Genomic features include 4410 CDSs, 102 RNAs, 3 CRISPR arrays, 3 prophage regions, and 37 predicted genomic islands. Two ß-lactone biosynthetic gene clusters were identified; besides, metabolic products of these are known to show antibiotic and/or anticancer properties. A siderophore biosynthetic gene cluster was also identified even though P. fragi is considered a non-siderophore producing pseudomonad. Other gene clusters of broad interest identified include those associated with bioremediation, biocontrol, plant growth promotion, or environmental adaptation. This dataset unveils various un-/underexplored metabolic or biosynthetic potential of P. fragi and provides insight into molecular mechanisms underpinning these attributes.

Synthesis, cytotoxicity and DNA binding of oxoazabenzo[de]anthracenes derivatives in colon cancer Caco-2 cells.

New oxoazabenzo[de]anthracenes derivatives were synthesised and characterised. Their interactions with calf thymus DNA were studied by UV spectrophotometric analysis and a competitive ethidium bromide displacement assay. Cytotoxicity was determined by MTT assay, against colon adenocarcinoma (Caco-2 cells). Among all the oxoazabenzo[de]anthracenes derivatives reported herein only the piperidino derivative exhibited strong DNA binding properties and cytotoxic activity with IC50 values in the range of 16 ± 1.5 µM (72-h treatment). In addition, the piperidino derivative did not directly inhibit topoisomerase I and topoisomerase II enzymes. The results confirm that the presence of the oxoazabenzo[de]anthracenes together with the piperidino functionality is crucial in exerting DNA binding and cytotoxic properties, hence demonstrating promise as a chemical scaffold for further development of new anticancer agents.

PEGylated derivatives of cystamine as enhanced treatments for nephropathic cystinosis.

The genetic disease, nephropathic cystinosis is characterized by lysosomal accumulation of the amino acid cystine. Crystallization of cystine in affected organs, if untreated, results in mortality of the affected individuals by their middle to late teens. The only approved treatment for cystinosis is administration of cysteamine. However, cysteamine is associated with an offending odor and taste and this, coupled to a rapid first pass metabolism and a 6h dosing regimen, suggest a clear need to improve the therapy. A number of PEGylated derivatives of cystamine, the disulfide counterpart of cysteamine, have been synthesised and evaluated in cultured cystinotic fibroblasts for toxicity and efficacy. All of the tested compounds were non-cytotoxic and displayed a remarkable depletion of intralysosomal cystine.

Oxoazabenzo[de]anthracenes conjugated to amino acids: synthesis and evaluation as DNA-binding antitumor agents.

We report the synthesis of an original series of oxoazabenzo[de]anthracenes conjugated to an amino acid: Ala, Phe, Pro, Lys, or Gly (4a-e, respectively). The compounds, derived from 1,8-dihydroxyanthracene-9,10-dione, were studied for DNA binding and cytotoxicity. Melting temperature, fluorescence quenching, and surface plasmon resonance methods all indicated that the lysine derivative 4d binds to DNA much more strongly that the Pro, Ala, and Gly conjugates whereas the Phe analogue showed the lowest DNA binding capacity. These compounds form intercalation complexes with DNA, as judged from electric linear dichroism and topoisomerase I-based DNA unwinding experiments. Preferential binding of 4d to defined sequences such as 5'-CTAAAGG and 5'-ATGC was evidenced by DNase I footprinting. This Lys conjugate was found to be over 20 times more cytotoxic to CEM human leukemia cells than the other conjugates, with an IC50 in the submicromolar range. A high antiproliferative activity, likely attributable to the enhanced DNA binding capacity, is maintained despite the incapacity of the compound to stabilize topoisomerase-DNA covalent complexes. The cell cycle effects of 4d consisted in an S phase accumulation of cells coupled with a pro-apoptotic action (appearance of hypodiploid sub-G1 cells) which were confirmed by measuring the inhibition of BrdU incorporation into DNA and labeling of phosphatidylserine residues with annexin V-FITC by means of flow cytometry. Altogether, the work provides interesting structure-activity relationships in the oxoazabenzo[de]anthracene-amino acid conjugate series and identifies the lysine derivative 4d as a promising candidate for further in vivo evaluation and drug design.