The over-expression of phasin and regulator genes promoting the synthesis of polyhydroxybutyrate in Cupriavidus necator H16 under non-stress conditions

Cupriavidus necator H16 is an ideal strain for polyhydroxybutyrate (PHB) production from CO 2 . Low-oxygen-stress can induce PHB synthesis in C. necator H16 while reducing bacterial growth under chemoautotrophic culture. The optimum growth and PHB synthesis of C. necator H16 cannot be achieved simultaneously, which restricts PHB production. The present study was initiated to address the issue through comparative transcriptome and gene function analysis. Firstly, the comparative transcriptome of C. necator H16 chemoautotrophically cultured under low-oxygen-stress and non-stress conditions was studied. Three types of transcription different genes were discovered: PHB enzymatic synthesis, PHB granulation, and regulators. Under low-oxygen-stress condition, acetoacetyl-CoA reductase gene phaB2 , PHB synthase gene phaC2 , phasins genes phaP1 and phaP2 , regulators genes uspA and rpoN were up-regulated 3.0, 2.5, 1.8, 2.7, 3.5, 1.6 folds, respectively. Secondly, the functions of up-regulated genes and their applications in PHB synthesis were further studied. It was found that the over-expression of phaP1 , phaP2 , uspA , and rpoN can induce PHB synthesis under non-stress condition, while phaB2 and phaC2 have no significant effect. Under the optimum condition, PHB percentage content in C. necator H16 was respectively increased by 37.2%, 28.4%, 15.8%, and 41.0% with the over-expression of phaP1 , phaP2 , uspA , and rpoN , and the corresponding PHB production increased by 49.8%, 42.9%, 47.0%, and 77.5% under non-stress chemoautotrophic conditions. Similar promotion by phaP1 , phaP2 , uspA , and rpoN was observed in heterotrophically cultured C. necator H16. The PHB percentage content and PHB production were respectively increased by 54.4% and 103.1% with the over-expression of rpoN under non-stress heterotrophic conditions. Importance Microbial fixation of CO 2 is an effective way to reduce greenhouse gases. Some microbes such as C. necator H16 usually accumulate PHB when they grow under stress. Low-oxygen-stress can induce PHB synthesis when C. necator H16 is autotrophically cultured with CO 2 , H 2 , and O 2 , while under stress, growth is restricted and total PHB yield is reduced. Achieving the optimal bacterial growth and PHB synthesis at the same time is an ideal condition for transforming CO 2 into PHB by C. necator H16. The present study was initiated to clarify the molecular basis of low-oxygen-stress promoting PHB accumulation and to realize the optimal PHB production by C. necator H16. Genes up-regulated under non-stress conditions were identified through comparative transcriptome analysis and over-expression of phasin and regulator genes were demonstrated to promote PHB synthesis in C. necator H16.

Dissolved oxygen as a propulsive parameter for polyhydroxyalkanoate production using Bacillus endophyticus cultures

Conventional biopolymers resembling synthetic polymers produced from microorganisms, polyhydroxyalkanoate (PHA) synthesized utilizing renewable resources have gained supreme attention recently. PHA accumulation within the microbial cell is an innate capability of bacteria to store carbon and energy when nutrient imbalance pertains. Gram positive Bacillus endophyticus capable of synthesizing PHA was focused in this study. Study focuses on the possibility of attaining high PHA yield in relation to the varying dissolved oxygen levels induced during production phase. There was a gradual increment in PHA production from 34.5 to 53.03% when cultivated in bioreactor that maintained least dissolved oxygen of 0.4 mg/L at 32 °C. The metabolic flux of organism was altered during oxygen stress brought by varying agitation rate and volume resulting in the accumulation of Nicotinamide Adenine Dinucleotide Hydrogen (NADH), which led to increase in the overall PHA production. PHA yield was found to be favored by decreasing the oxygen supply thereby inducing an oxygen stress environment. This report was the first one that was correlating the hypothesis that links PHA yield and oxygen stress condition during production phase. PHA produced was characterized by FTIR and 1HNMR spectra in which the presence of Polyhydroxybutyrate was confirmed. Graphical abstract

Application of Dynamic Controlled Atmosphere Technologies to Reduce Incidence of Physiological Disorders and Maintain Quality of ‘Granny Smith’ Apples

The efficacy of dynamic controlled atmosphere technologies; repeated low oxygen stress (RLOS) and dynamic controlled atmosphere-chlorophyll fluorescence (DCA-CF) to control superficial scald development on ‘Granny Smith’ apples during long-term storage was studied. Fruit were stored for 2, 4, 6, 8, and 10 months at 0 °C in DCA-CF (0.6% O2 and 0.8% CO2), regular atmosphere (RA)(≈21% O2 and 90–95% RH), and RLOS treatments: (1) 0.5% O2 for 10 d followed by ultra-low oxygen (ULO) (0.9% O2 and 0.8% CO2) for 21 d and 0.5% O2 for 7 d or (2) 0.5% O2 for 10 d followed by controlled atmosphere (CA) (1.5% O2 and 1% CO2) for 21 d and 0.5% O2 for 7 d. Development of superficial scald was inhibited for up to 10 months and 7 d shelf life (20 °C) under RLOS + ULO and DCA-CF treatments. Apples stored in RLOS + ULO, RLOS + CA, and DCA-CF had significantly (p < 0.05) higher flesh firmness and total soluble solids. The RLOS phases applied with CA or ULO and DCA-CF storage reduced the development of superficial scald by possibly suppressing the oxidation of volatiles implicated in superficial scald development.

Pyrophosphate as an alternative energy currency in plants

In the conditions of [Mg2+] elevation that occur, in particular, under low oxygen stress and are the consequence of the decrease in [ATP] and increase in [ADP] and [AMP], pyrophosphate (PPi) can function as an alternative energy currency in plant cells. In addition to its production by various metabolic pathways, PPi can be synthesized in the combined reactions of pyruvate, phosphate dikinase (PPDK) and pyruvate kinase (PK) by so-called PK/PPDK substrate cycle, and in the reverse reaction of membrane-bound H+-pyrophosphatase, which uses the energy of electrochemical gradients generated on tonoplast and plasma membrane. The PPi can then be consumed in its active forms of MgPPi and Mg2PPi by PPi-utilizing enzymes, which require an elevated [Mg2+]. This ensures a continuous operation of glycolysis in the conditions of suppressed ATP synthesis, keeping metabolism energy efficient and less dependent on ATP.

The Oxidative Paradox in Low Oxygen Stress in Plants

Reactive oxygen species (ROS) are part of aerobic environments, and variations in the availability of oxygen (O2) in the environment can lead to altered ROS levels. In plants, the O2 sensing machinery guides the molecular response to low O2, regulating a subset of genes involved in metabolic adaptations to hypoxia, including proteins involved in ROS homeostasis and acclimation. In addition, nitric oxide (NO) participates in signaling events that modulate the low O2 stress response. In this review, we summarize recent findings that highlight the roles of ROS and NO under environmentally or developmentally defined low O2 conditions. We conclude that ROS and NO are emerging regulators during low O2 signalling and key molecules in plant adaptation to flooding conditions.