Person: TOKSOY ÖNER, EBRU
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TOKSOY ÖNER
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Publication Metadata only The Stimulatory Effect of Mannitol on Levan Biosynthesis: Lessons from Metabolic Systems Analysis of Halomonas smyrnensis AAD6(T)(WILEY, 2013) TOKSOY ÖNER, EBRU; Ates, Ozlem; Arga, Kazim Y.; Oner, Ebru ToksoyHalomonas smyrnensis AAD(T) is a halophilic, gram-negative bacterium that can efficiently produce levan from sucrose as carbon source via levansucrase activity. However, systems-based approaches are required to further enhance its metabolic performance for industrial application. As an important step toward this goal, the genome-scale metabolic network of Chromohalobacter salexigens DSM3043, which is considered a model organism for halophilic bacteria, has been reconstructed based on its genome annotation, physiological information, and biochemical information. In the present work, the genome-scale metabolic network of C. salexigens was recruited, and refined via integration of the available biochemical, physiological, and phenotypic features of H. smyrnensis AAD6(T). The generic metabolic model, which comprises 1,393 metabolites and 1,108 reactions, was then systematically analyzed in silico using constraints-based simulations. To elucidate the relationship between levan biosynthesis and other metabolic processes, an enzyme-graph representation of the metabolic network and a graph decomposition technique were employed. Using the concept of control effective fluxes, significant links between several metabolic processes and levan biosynthesis were estimated. The major finding was the elucidation of the stimulatory effect of mannitol on levan biosynthesis, which was further verified experimentally via supplementation of mannitol to the fermentation medium. The optimal concentration of 30 g/L mannitol supplemented to the 50 g/L sucrose-based medium resulted in a twofold increase in levan production in parallel with increased sucrose hydrolysis rate, accumulated extracellular glucose, and decreased fructose uptake rate. (c) 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:1386-1397, 2013Publication Metadata only Sulfated levan from Halomonas smyrnensis as a bioactive, heparin-mimetic glycan for cardiac tissue engineering applications(ELSEVIER SCI LTD, 2016) TOKSOY ÖNER, EBRU; Erginer, Merve; Akcay, Ayca; Coskunkan, Binnaz; Morova, Tunc; Rende, Deniz; Bucak, Seyda; Baysal, Nihat; Ozisik, Rahmi; Eroglu, Mehmet S.; Agirbasli, Mehmet; Oner, Ebru ToksoyChemical derivatives of levan from Halomonas smyrnensis AAD6T with low, medium and high levels of sulfation were synthesized and characterized by FTIR and 2D-NMR. Sulfated levan samples were found to exhibit anticoagulation activity via the intrinsic pathway like heparin in a dose-dependent manner. Exceptionally high heparin equivalent activity of levan sulfate was shown to proceed via thrombin inhibition where decreased Factor Xa activity with increasing concentration was observed in antithrombin tests and above a certain concentration, levan sulfate showed a better inhibitor activity than heparin. In vitro experimental results were then verified in silico by docking studies using equilibrium structures obtained by molecular dynamic simulations and results suggested a sulfation dependent binding mechanism. With its high biocompatibility and heparin mimetic activity, levan sulfate can be considered as a suitable functional biomaterial to design biologically active, functionalized, thin films and engineered smart scaffolds for cardiac tissue engineering applications. (C) 2016 Elsevier Ltd. All rights reserved.Publication Metadata only Effective stimulating factors for microbial levan production by Halomonas smyrnensis AAD6(T)(SOC BIOSCIENCE BIOENGINEERING JAPAN, 2015) TOKSOY ÖNER, EBRU; Sarilmiser, Hande Kazak; Ates, Ozlem; Ozdemir, Gonca; Arga, Kazim Yalcin; Oner, Ebru ToksoyLevan is a bioactive fructan polymer that is mainly associated with high-value applications where exceptionally high purity requirements call for well-defined cultivation conditions. In this study, microbial levan production by the halophilic extremophile Halomonas smyrnensis AAD6(T) was investigated systematically. For this, different feeding strategies in fed-batch cultures were employed and fermentation profiles of both shaking and bioreactor cultures were analyzed. Initial carbon and nitrogen source concentrations, production pH, NaCl and nitrogen pulses, nitrogen and phosphorous limitations, trace elements and thiamine contents of the basal production medium were found to affect the levan yields at different extends. Boric acid was found to be the most effective stimulator of levan production by increasing the sucrose utilization three-fold and levan production up to five-fold. This significant improvement implied the important role of quorum sensing phenomenon and its regulatory impact on levan production mechanism. Levan produced by bioreactor cultures under conditions optimized within this study was found to retain its chemical structure. Moreover, its biocompatibility was assessed for a broad concentration range. Hence H. smyrnensis AAD6(T) has been firmly established as an industrially important resource microorganism for high-quality levan production. (C) 2014, The Society for Biotechnology, Japan. All rights reserved.Publication Metadata only The Genome-Based Metabolic Systems Engineering to Boost Levan Production in a Halophilic Bacterial Model(MARY ANN LIEBERT, INC, 2018) TOKSOY ÖNER, EBRU; Aydin, Busra; Ozer, Tugba; Oner, Ebru Toksoy; Arga, Kazim YalcinMetabolic systems engineering is being used to redirect microbial metabolism for the overproduction of chemicals of interest with the aim of transforming microbial hosts into cellular factories. In this study, a genome-based metabolic systems engineering approach was designed and performed to improve biopolymer biosynthesis capability of a moderately halophilic bacterium Halomonas smyrnensis AAD6(T) producing levan, which is a fructose homopolymer with many potential uses in various industries and medicine. For this purpose, the genome-scale metabolic model for AAD6(T) was used to characterize the metabolic resource allocation, specifically to design metabolic engineering strategies for engineered bacteria with enhanced levan production capability. Simulations were performed in silico to determine optimal gene knockout strategies to develop new strains with enhanced levan production capability. The majority of the gene knockout strategies emphasized the vital role of the fructose uptake mechanism, and pointed out the fructose-specific phosphotransferase system (PTSfru) as the most promising target for further metabolic engineering studies. Therefore, the PTSfru of AAD6(T) was restructured with insertional mutagenesis and triparental mating techniques to construct a novel, engineered H. smyrnensis strain, BMA14. Fermentation experiments were carried out to demonstrate the high efficiency of the mutant strain BMA14 in terms of final levan concentration, sucrose consumption rate, and sucrose conversion efficiency, when compared to the AAD6(T). The genome-based metabolic systems engineering approach presented in this study might be considered an efficient framework to redirect microbial metabolism for the overproduction of chemicals of interest, and the novel strain BMA14 might be considered a potential microbial cell factory for further studies aimed to design levan production processes with lower production costs.Publication Metadata only Halomonas smyrnensis as a cell factory for co-production of PHB and levan(ELSEVIER SCIENCE BV, 2018) KASAVİ, CEYDA; Tohme, Souha; Haciosmanoglu, Gul Gulenay; Eroglu, Mehmet Sayip; Kasavi, Ceyda; Genc, Seval; Can, Zehra Semra; Oner, Ebru ToksoyLevan is a fructan type polysaccharide that has long been considered as an industrially important biopolymer however its limited availability is mainly due to the bottlenecks associated with its large-scale production. To overcome such bottlenecks in the commercialization of this very promising polysaccharide, co-production of levan with polyhydroxyalkanoates (PHAs) by halophilic Halomonas smyrnensis cultures has been proposed in this study for the first time. After in silico and in vitro assessment of PHA accumulation, fermentation profiles for levan and PHA concentrations were obtained in the presence of sucrose and glucose and the PHA granules observed by TEM were found to be poly(3-hydroxybutyrate) (PHB) after detailed structural characterization by GC-MS, DSC, FTIR and NMR. Six nutrient limitation strategies based on nitrogen (N) and phosphorus (P) were tested but highest levan and PHB yields were obtained under unlimited conditions. H. smyrnensis is proved to co-produce PHB and levan while using inexpensive carbon sources which is a commercially successful microbial cell factory system showing a great potential in lowering manufacturing costs and aiming for a zero waste policy within the biorefinery concept. (C) 2018 Elsevier B.V. All rights reserved.Publication Metadata only Identification of N-Hexadecanoyl-L-homoserine lactone (C16-AHL) as signal molecule in halophilic bacterium Halomonas smyrnensis AAD6(SPRINGER, 2016) TOKSOY ÖNER, EBRU; Abbamondi, Gennaro Roberto; Suner, Salim; Cutignano, Adele; Grauso, Laura; Nicolaus, Barbara; Oner, Ebru Toksoy; Tommonaro, GiuseppinaQuorum sensing (QS) mechanisms regulate a variety of vital functions such as motility, growth inhibition, biofilm development, virulence expression and plasmid conjugation. Many of these activities are essential for the well-being of organisms, in particular in extreme conditions such as hypersaline environments. Halophiles are able to synthesize products that are stable, and to exploit their activities under such extreme conditions, namely saline environments such as marine habitat, salt lakes, brines and saline soils. The versatility of these products leads researchers to address their interest in biotechnological applications. In this study, the QS activity of Halomonas smyrnensis AAD6-a moderately halophilic and exopolysaccharide-producing bacterium-was investigated. Dichloromethane extracts from stationary phase cultures of Halomonas smyrnensis AAD6 were partially purified, and C-16-AHL was identified by LC-MS analysis. The synthesis of autoinducers was assayed and their production was detected starting from 48 h of growth. The growth-phase dependent production of exopolysaccharide (EPS) and signal molecules suggested that these activities could be linked.Publication Metadata only Development of a cost-effective production process for Halomonas levan(SPRINGER, 2018) TOKSOY ÖNER, EBRU; Erkorkmaz, Burak Adnan; Kirtel, Onur; Duru, Ozlem Ates; Oner, Ebru ToksoyLevan polysaccharide is an industrially important natural polymer with unique properties and diverse high-value applications. However, current bottlenecks associated with its large-scale production need to be overcome by innovative approaches leading to economically viable processes. Besides many mesophilic levan producers, halophilic Halomonas smyrnensis cultures hold distinctive industrial potential and, for the first time with this study, the advantage of halophilicity is used and conditions for non-sterile levan production were optimized. Levan productivity of Halomonas cultures in medium containing industrial sucrose from sugar beet and food industry by-product syrup, a total of ten sea, lake and rock salt samples from four natural salterns, as well as three different industrial-grade boron compounds were compared and the most suitable low-cost substitutes for sucrose, salt and boron were specified. Then, the effects of pH control, non-sterile conditions and different bioreactor modes (batch and fed-batch) were investigated. The development of a cost-effective production process was achieved with the highest yield (18.06 g/L) reported so far on this microbial system, as well as the highest theoretical bioconversion efficiency ever reported for levan-producing suspension cultures. Structural integrity and biocompatibility of the final product were also verified in vitro.