Previously we constructed a Bacillus subtilis strain for efficient production of N-acetylglucosamine (GlcNAc) by engineering of GlcNAc synthetic and catabolic pathways. However, the further improvement of GlcNAc titer is limited by the intrinsic inefficiency of GlcNAc synthetic pathway and undesirable cellular properties including sporulation and high maintenance metabolism. In this work, we further improved GlcNAc titer through spatial modulation of key pathway enzymes and by blocking sporulation and decreasing maintenance metabolism. Specifically, a DNA-guided scaffold system was firstly used to modulate the activities of glucosamine-6-phosphate synthase and GlcNAc-6-phosphate N-acetyltransferase, increasing the GlcNAc titer from 1.83 g/L to 4.55 g/L in a shake flask. Next, sporulation was blocked by respectively deleting spo0A (gene encoding the initiation regulon of sporulation) and sigE (gene encoding RNA polymerase sporulation-specific sigma factor). Deletion of sigE more effectively blocked sporulation without altering cell growth or GlcNAc production. The respiration chain was then engineered to decrease the maintenance metabolism of recombinant B. subtilis by deleting cydB and cydC, genes encoding cytochrome bd ubiquinol oxidase (subunit II) and ATP-binding protein for the expression of cytochrome bd, respectively. The respiration-engineered B. subtilis produced 6.15 g/L GlcNAc in a shake flask and 20.58 g/L GlcNAc in a 3-L fed-batch bioreactor. To the best of our knowledge, this report is the first to describe the modulation of pathway enzymes via a DNA-guided scaffold system in B. subtilis. The combination of spatial modulation of key pathway enzymes and optimization of cellular properties may be used to develop B. subtilis as a well-organized cell factory for the production of the other industrially useful chemicals. 相似文献
Abstract: The retentostat was developed for long-term continuous, axenic cultivation of microorganisms at those low growth rates which prevail in most natural habitats and which cannot be established properly in chemostats. How a microbial population approaches 'zero-growth' was studied in axenic cultures of Nitrosomonas europaea with complete biomass retention at 25°C and constant input of a nutrient solution containing ammonium (0.57 mM) as energy source. Since only cell-free filtrate left the reactor, biomass accumulated until a stable maximum of 2.7 × 109 cells ml−1 (398 mg l−1 dry matter) was reached after about 5 weeks. In this state, growth rate approached zero, and the ammonium input just met the substrate demand required for maintenance energy (1.43 μmol NH3–N mg dm−1 h−1). The potential of the retentostat for studying interactions between different microorganisms was demonstrated with a cascade of cultures of Nitrosomonas, Nitrobacter , and a denitrifying Pseudomonas . Thereby the ammonia was completely eliminated from artificial wastewater. 相似文献
Twenty four Awassi lambs (mean BW 24.4 kg) were used in a two periods experiment to measure maintenance (zero growth) and growth requirements of metabolizable energy (ME). During the pre-treatment period (period 1, 3 weeks) all lambs were maintained at a constant level of feed intake (2% BW). During the treatment period (period 2, 10 weeks) the lambs were divided into four equal groups and fed different amounts (2, 3, 4 and 5% of BW). Six additional lambs of similar BW and age were killed at the beginning of the experiment to compare final empty body weight (FEBW) with initial body weight (IBW). Twelve lambs, divided into four equal groups were fed the experimental diet in the same amounts as in the growth experiment (period 2) to determine the digestibility of energy.
Energy requirement for maintenance (EM) was measured by both constant weight technique, using data collected during period 1, and regression technique by regressing ME intake on body weight gain (BWG) and empty body weight gain (EBWG). EM measured during the constant weight period (pre-treatment) and overall experimental period (pre-treatment plus treatment periods) were 0.482 and 0.466 MJ of ME per kg M0.75 per day. Predicted energy requirements for growth (Eg) calculated from equations derived during the treatment period were 0.433, 0.623, 0.782, 0.910 and 1.007 (MJ per kg M0.75 per day) for the growth rates 50, 100, 150, 200, and 250 (g per day) respectively. It is concluded to be more appropriate to use the values derived during the overall experimental period for maintenance and the overall treatment period for growth as they are most applicable to production situations. 相似文献
Zymomonas mobilis growing aerobically with 20 g glucose–1 (carbon-limited) in a chemostat exhibited an increase in both the molar growth yield (Yx/s) and the maximum molar growth yield (Yx/smax) and a decrease in both the specific substrate consumption rate (qs) and the maintenance energy consumption rate (me). Stepwise increase in the input oxygen partial pressure showed that anaerobic-to-aerobic transitional adaptation occurred
in four stages: anaerobic (0 mm HgO2), oxygen-limited (7.6– 230 mm HgO2), intermediate (273 mm HgO2), and oxygen excess (290 mm HgO2). The steady-state biomass concentration, Yx/s, and intracellular ATP content increased between oxygen partial pressures of 7.6 and 120 mm HgO2, accompanied by a decrease in the qs and the specific acid production rate. The membrane ATPase activity decreased with increasing oxygen partial pressure and
reached its lowest levels at 273 mm HgO2, which was the highest input oxygen partial pressure where steady-state conditions were possible. Glucokinase, glucose-6-phosphate
dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase, and alcohol dehydrogenase activities also decreased when the oxygen
partial pressure was increased above 15 mm Hg, whereas pyruvate decarboxylase was unaffected by aeration. Growth inhibition
at 290 mm HgO2 was characterised by a drastic reduction in the pyruvate kinase activity and a collapse in the intracellular ATP pool. The
growth and enzyme data suggest that at low glucose concentrations and oxygen-limited conditions, the increase in biomass yields
is a reflection of a redirection of ATP usage rather than a net increase in energy production.
Received: 14 August 1996 / Accepted: 31 January 1997 相似文献
The ecosystems of Chernobyl nuclear accident zone, having been deeply transformed by normal long-term agricultural and forestry activity, have been damaged by irradiation after the nuclear accident of 1986 and then influenced by post-accident countermeasures. Ecosystem remediation is being undertaken in this area, including maintenance of the forests, artificial afforestation, and processes of spontaneous self-restoration of the natural coenoses. Both natural and artificial remediation of the landscapes and ecosystems at the abandoned territory improves their capacity to retain radionuclides and their ability to stabilise the processes of redistribution of radionuclides and their spread in the environment. The Chernobyl zone is a territory of great importance for studying ecosystem remediation processes and elaborating on ecological engineering technologies to correct serious environmental problems arising from radionuclides release. 相似文献
目的:探讨血液透析前后和血液透析过程中血压及其变异性与患者预后的相关性。方法:选取沈阳军区总医院血液透析中心2010年1月1日至2013年12月31日收治的维持性血液透析患者,收集并对比随访期内生存及死亡患者的自然信息及临床资料,评估血液透析过程中患者血压及其变异程度与患者预后的关系。结果:共有269例患者纳入研究,经过4年随访共死亡83(30.86%)例。死亡组年龄显著大于存活组(64.92±13.24岁比49.89±12.86岁,P=0.000),而透析年限显著短于存活组(2.60±2.56年比7.25±4.14年,P=0.000)。死亡组透析前SBP(P=0.001)、DBP(P=0.000)、MAP(P=0.000)均显著高于存活组。两组间透析后SBP、DBP、MAP比较均无显著差异。死亡组△SBP(P=0.026)、△DBP(P=0.001)、△MAP(P=0.001)幅度显著大于存活组。死亡组透析前SBP变异率显著高于存活组(P=0.001);死亡组透析后SBP变异率(P=0.000)、DBP变异率(P=0.014)、MAP变异率(P=0.005)均高于存活组。死亡组每次透析前各时间点间SBP变异率(0.12±0.04 mm Hg比0.09±0.03 mm Hg,P=0.000)与MAP变异率(0.10±0.03mm Hg比0.09±0.03 mm Hg,P=0.001)显著高于非死亡组。结论:维持性血液透析患者透析前血压、透析前后血压改变幅度、透析前后血压变异率、每次透析各时间点间血压变异率等与全因死亡相关。 相似文献