Biotic inactivation of the Pseudomonas aeruginosa quinolone signal molecule

In Pseudomonas aeruginosa, quorum sensing (QS) regulates the production of secondary metabolites, many of which are antimicrobials that impact on polymicrobial community composition. Consequently, quenching QS modulates the environmental impact of P. aeruginosa. To identify bacteria capable of inactivating the QS signal molecule 2-heptyl-3- hydroxy-4(1H)-quinolone (PQS), a minimal medium containing PQS as the sole carbon source was used to enrich a Malaysian rainforest soil sample. This yielded an Achromobacter xylosoxidans strain (Q19) that inactivated PQS, yielding a new fluorescent compound (I-PQS) confirmed as PQS-derived using deuterated PQS. The I-PQS structure was elucidated using mass spectrometry and nuclear magnetic resonance spectroscopy as 2-heptyl-2-hydroxy-1,2-dihydroquinoline- 3,4-dione (HHQD). Achromobacter xylosoxidans Q19 oxidized PQS congeners with alkyl chains ranging from C1 to C5 and also N-methyl PQS, yielding the corresponding 2-hydroxy-1,2-dihydroquinoline-3,4- diones, but was unable to inactivate thePQSprecursor HHQ. This indicates that the hydroxyl group at position 3 in PQS is essential and that A. xylosoxidans inactivates PQS via a pathway involving the incorporation of oxygen at C2 of the heterocyclic ring. The conversion of PQS to HHQD also occurred on incubation with 12/17 A. xylosoxidans strains recovered from cystic fibrosis patients, with P. aeruginosa and with Arthrobacter, suggesting that formation of hydroxylated PQS may be a common mechanism of inactivation

Biotic inactivation of the Pseudomonas aeruginosa quinolone signal molecule

In Pseudomonas aeruginosa, quorum sensing (QS) regulates the production of secondary metabolites, many of which are antimicrobials that impact on polymicrobial community composition. Consequently, quenching QS modulates the environmental impact of P. aeruginosa. To identify bacteria capable of inactivating the QS signal molecule 2-heptyl-3- hydroxy-4(1H)-quinolone (PQS), a minimal medium containing PQS as the sole carbon source was used to enrich a Malaysian rainforest soil sample. This yielded an Achromobacter xylosoxidans strain (Q19) that inactivated PQS, yielding a new fluorescent compound (I-PQS) confirmed as PQS-derived using deuterated PQS. The I-PQS structure was elucidated using mass spectrometry and nuclear magnetic resonance spectroscopy as 2-heptyl-2-hydroxy-1,2-dihydroquinoline- 3,4-dione (HHQD). Achromobacter xylosoxidans Q19 oxidized PQS congeners with alkyl chains ranging from C1 to C5 and also N-methyl PQS, yielding the corresponding 2-hydroxy-1,2-dihydroquinoline-3,4- diones, but was unable to inactivate thePQSprecursor HHQ. This indicates that the hydroxyl group at position 3 in PQS is essential and that A. xylosoxidans inactivates PQS via a pathway involving the incorporation of oxygen at C2 of the heterocyclic ring. The conversion of PQS to HHQD also occurred on incubation with 12/17 A. xylosoxidans strains recovered from cystic fibrosis patients, with P. aeruginosa and with Arthrobacter, suggesting that formation of hydroxylated PQS may be a common mechanism of inactivation

Structure and function of negative feedback loops at the interface of genetic and metabolic networks

The molecular network in an organism consists of transcription/translation regulation, protein-protein interactions/modifications and a metabolic network, together forming a system that allows the cell to respond sensibly to the multiple signal molecules that exist in its environment. A key part of this overall system of molecular regulation is therefore the interface between the genetic and the metabolic network. A motif that occurs very often at this interface is a negative feedback loop used to regulate the level of the signal molecules. In this work we use mathematical models to investigate the steady state and dynamical behaviour of different negative feedback loops. We show, in particular, that feedback loops where the signal molecule does not cause the dissociation of the transcription factor from the DNA respond faster than loops where the molecule acts by sequestering transcription factors off the DNA. We use three examples, the bet, mer and lac systems in E. coli, to illustrate the behaviour of such feedback loops.Comment: 8 pages, 4 figure

Chemical communication between synthetic and natural cells: a possible experimental design

The bottom-up construction of synthetic cells is one of the most intriguing and interesting research arenas in synthetic biology. Synthetic cells are built by encapsulating biomolecules inside lipid vesicles (liposomes), allowing the synthesis of one or more functional proteins. Thanks to the in situ synthesized proteins, synthetic cells become able to perform several biomolecular functions, which can be exploited for a large variety of applications. This paves the way to several advanced uses of synthetic cells in basic science and biotechnology, thanks to their versatility, modularity, biocompatibility, and programmability. In the previous WIVACE (2012) we presented the state-of-the-art of semi-synthetic minimal cell (SSMC) technology and introduced, for the first time, the idea of chemical communication between synthetic cells and natural cells. The development of a proper synthetic communication protocol should be seen as a tool for the nascent field of bio/chemical-based Information and Communication Technologies (bio-chem-ICTs) and ultimately aimed at building soft-wet-micro-robots. In this contribution (WIVACE, 2013) we present a blueprint for realizing this project, and show some preliminary experimental results. We firstly discuss how our research goal (based on the natural capabilities of biological systems to manipulate chemical signals) finds a proper place in the current scientific and technological contexts. Then, we shortly comment on the experimental approaches from the viewpoints of (i) synthetic cell construction, and (ii) bioengineering of microorganisms, providing up-to-date results from our laboratory. Finally, we shortly discuss how autopoiesis can be used as a theoretical framework for defining synthetic minimal life, minimal cognition, and as bridge between synthetic biology and artificial intelligence.Comment: In Proceedings Wivace 2013, arXiv:1309.712

Leucine as a signal molecule

Biosignalizacijom se stanice (tkiva kod višestaničnih organizama) prilagođavaju izvanstaničnim uvjetima (staničnom okruženju) i na posredan način komuniciraju s okolinom tako da aktiviraju ili utišaju ekspresiju ciljanih gena, a time i metaboličke puteve. Signalizacija se temelji na pretvorbi primljenog izvanstaničnog signala u unutarstanični signal koji potiče ili utišava ciljane stanične procese. Prijenos signala uključuje proces umnažanja signala i prevođenja jedne vrste signala u drugi signal koja stanici predstavlja određenu informaciju. Jedna od signalnih molekula je i aminokiselina L-leucin. Molekula L-leucina je signal koji označava uvjete energetskog obilja okoline,a prenosi se u stanicu transporterom LAT1. Cilj ovog rada bio je opisati L-leucin kao signalnu molekulu koja djeluje na transkripciju gena te metabolizam. Regulatorni protein ovisan o leucinu, Lrp, i protein Sestrin2 su unutarstanični senzori leucina, a vezanjem leucina su potaknuti na konformacijsku promjenu. Lrp, protein iz bakterije Escherichia coli, je transkripcijski regulator koji prenosi informaciju leucina na molekulu DNA čime se aktivira/utiša transkripcija gena. Na ovaj način L-leucin djeluje na transkripciju gena tRNA, rRNA, katabolizma aminokiselina, fimbrija i transkripcijskih faktora. Sestrin2 je citosolni protein sisavaca koji prenosi singal L-leucina preko posredničkih proteina do krajnjeg akceptora signala - proteinskog kompleksa mTORC1. mTORC1 je arhitektura koji sadrži protein-kinazu mTOR koja fosforilacijom ciljanih proteina potiče ili inhibira njihovu aktivnost. Fosforilirani proteini sudjeluju u sintetskim reakcijama proteina i lipida, metabolizmu koji sudjeluje u pretvorbi energije i autofagiji. Na taj način L-leucin djeluje na metaboličke puteve stanice.Biosignalization helps cells to adapt to the extracellular conditions (environment) and indirectly communicate with environment by activating/silencing expression of target genes and metabolism, respectively. Signalization is based on conversion of extracellular signal to the intracellular signal which activates or inhibits many cellular processes. Transduction of the signal includes the process of signal amplification and signal translation from one type to another. After the signal has been interpreted, it provides a cell a specific information. One of the signal molecules is an amino acid L-leucine. The aim od this report was to present L-leucine as a signal molecule for gene transcription and metabolism. L-leucine is an signal that indicates nutrient-rich conditions and is transported into the cell by LAT1 transporter. Leucine-responsive regulatory protein, Lrp, and protein Sestrin2 are intracellular L-leucine sensors that are induced to conformational change by binding of leucine. Lrp, a Escherichia coli protein, is a transcriptional regulator which transduces L-leucine information to the DNA and therefore activates/represses transcription of target genes. In this way, L-leucine effects transcription of tRNA, rRNA, amino acid catabolism genes, fimbria and transcription factors genes. Sestrin2 is a mammalian cytosolic protein that transfers L-leucine signal through mediator proteins to the terminal signal acceptor – protein complex mTORC1. mTORC1 is an architecture that includes mTOR protein kinase. Once mTOR has phosphorilated protein, actvity of a protein is activated or repressed. Phosphorilated proteins take part in lipid and protein biosynthesis reactions, energy metabolism and autophagy. In this way, L-leucine acts on the metabolic pathways of the cell

Myocardial Protective Effect of Gas Signal Molecule Hydrogen Sulfide on Cardiovascular Disease

Cardiovascular diseases increase continually in the worldwide scale, and its specific pathogenesis has not been completely clear. The gas signal molecule hydrogen sulfide (H2S) is a new type of neuroactive substance, which plays many biological roles in many systems such as cardiovascular system. In recent years, a lot of research has confirmed H2S has myocardial protective effect on cardiovascular diseases such as atherosclerosis, ischemia-reperfusion injury, hypertension and heart failure. This paper reviews the research status of myocardial protective effect of H2S on cardiovascular diseases