RESUMO
Ligand-induced system plays an important role for microbial engineering due to its tunable gene expression control over timings and levels. An oleic acid (OA)-induced system was recently constructed based on protein FadR, a transcriptional regulator involved in fatty acids metabolism, for metabolic control in Escherichia coli. In this study, we constructed a synthetic FadR-based OA-induced systems in Halomonas bluephagenesis by hybridizing the porin promoter core region and FadR-binding operator (fadO). The dynamic control range was optimized over 150-fold, and expression leakage was significantly reduced by tuning FadR expression and positioning fadO, forming a series of OA-induced systems with various expression strengths, respectively. Additionally, ligand orthogonality and cross-species portability were also studied and showed highly linear correlation among Halomonas spp., Escherichia coli and Pseudomonas spp. Finally, OA-induced systems with medium- and small-dynamic control ranges were employed to dynamically control the expression levels of morphology associated gene minCD, and monomer precursor 4-hydroxybutyrate-CoA (4HB-CoA) synthesis pathway for polyhydroxyalkanoates (PHA), respectively, in the presence of oleic acid as an inducer. As a result, over 10 g/L of poly-3-hydroxybutyrate (PHB) accumulated by elongated cell sizes, and 6 g/L of P(3HB-co-9.57 mol% 4HB) were obtained by controlling the dose and induction time of oleic acid only. This study provides a systematic approach for ligand-induced system engineering, and demonstrates an alternative genetic tool for dynamic control of industrial biotechnology.
Assuntos
Halomonas , Poli-Hidroxialcanoatos , Coenzima A/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Halomonas/genética , Halomonas/metabolismo , Ligantes , Engenharia Metabólica , Ácido Oleico/metabolismo , Poliésteres/metabolismo , Poli-Hidroxialcanoatos/genética , Pseudomonas/genética , Pseudomonas/metabolismoRESUMO
Cross-linking is known to play a pivotal role in the relaxation dynamics and mechanical properties of thermoset polymers, which are commonly used in structural applications because of their light weight and inherently strong nature. Here, we employ a coarse-grained (CG) polymer model to systematically explore the effect of cross-link density on basic thermodynamic properties as well as corresponding changes in the segmental dynamics and elastic properties of these network materials upon approaching their glass transition temperatures (Tg). Increasing the cross-link density unsurprisingly leads to a significant slowing down of the segmental dynamics, and the fragility K of glass formation shifts in lockstep with Tg, as often found in linear polymer melts when the polymer mass is varied. As a consequence, the segmental relaxation time τα becomes almost a universal function of reduced temperature, (T - Tg)/Tg, a phenomenon that underlies the applicability of the "universal" Williams-Landel-Ferry (WLF) relation to many polymer materials. We also test a mathematical model of the temperature dependence of the linear elastic moduli based on a simple rigidity percolation theory and quantify the fluctuations in the local stiffness of the network material. The moduli and distribution of the local stiffness likewise exhibit a universal scaling behavior for materials having different cross-link densities but fixed (T - Tg)/Tg. Evidently, Tg dominates both τα and the mechanical properties of our model cross-linked polymer materials. Our work provides physical insights into how the cross-link density affects glass formation, aiding in the design of cross-linked thermosets and other structurally complex glass-forming materials.
RESUMO
Thermoset materials often involve the addition of molecular and nanoparticle additives to alter various chemo-physical properties of importance in their ultimate applications. The resulting compositional heterogeneities can lead to either enhancement or degradation of thermoset properties, depending on the additive chemical structure and concentration. We tentatively explore this complex physical phenomenon through the consideration of a model polymeric additive to our coarse-grained (CG) thermoset investigated in previous works by simply varying the size of additive segments compared to those of polymer melt. We find that the additive modified thermoset material becomes chemically heterogeneous from additive aggregation when the additive segments become much smaller than those of the thermoset molecules, and a clear evidence is observed in the spatial distribution of local molecular stiffness estimated from Debye-Waller factor ãu2ã. Despite the non-monotonic variation trends observed in dynamical and mechanical properties with decreasing additive segmental size, both the structural relaxation time and moduli (i.e., shear modulus and bulk modulus) exhibit scaling laws with ãu2ã. The present work highlights the complex role of additive size played in the dynamical and mechanical properties of thermoset polymers, which should provide a better understanding for the glass formation process of cross-linked polymer composites.