Enhanced extracellular production of alpha-lactalbumin from Bacillus subtilis through signal peptide and promoter screening

Alpha-lactalbumin(α-LA)is a major whey protein found in breast milk and plays a crucial role in the growth and development of infants.In this study,Bacillus subtilis RIK1285 harboring AprE signal peptide(SP)was selected as the original strain for the production ofα-LA.It was found thatα-LA was identified in the pellet after ultrasonic disruption and centrifugation instead of in the fermentation supernatant.The original strain most likely only producedα-LA intracellular,but not extracellular.To improve the expression and secretion ofα-LA in RIK1285,a library of 173 homologous SPs from the B.subtilis 168 genome was fused with target LALBA gene in the pBE-S vector and expressed extracellularly in RIK1285.SP YjcN was determined to be the best signal peptide.Bands in supernatant were observed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and purified by nickel column to calculate the highest yield signal peptide.In addition,different promoters(P_(aprE),P_(43),and P_(glv))were compared and applied.The results indicated that the strain RIK1285-pBE-P_(glv)-YjcN-LALBA had the highestα-LA yield,reaching 122.04μg/mL.This study demonstrates successful expression and secretion of humanα-LA in B.subtilis and establishes a foundation for simulating breast milk for infant formulas and developing bioengineered milk.

Remodeling Isoprene Pyrophosphate Metabolism for Promoting Terpenoids Bioproduction

Terpenoids are the largest family of natural products.They are made from the building block isoprene pyrophosphate(IPP),and their bioproduction using engineered cell factories has received a great deal of attention.To date,the insufficient metabolic supply of IPP remains a great challenge for the efficient synthesis of terpenoids.In this work,we discover that the imbalanced metabolic flux distribution between the central metabolism and the IPP supply hinders IPP accumulation in Bacillus subtilis(B.subtilis).Therefore,we remodel the IPP metabolism using a series of genetically encoded two-input-multioutput(TIMO)circuits that are responsive to pyruvate or/and malonyl-CoA,resulting in an IPP pool that is significantly increased by up to four-fold.As a proof-of-concept validation,we design an IPP metabolism remodeling strategy to improve the production of three valuable terpenoids,including menaquinone-7(MK-7,4.1-fold),lycopene(9-fold),andβ-carotene(0.9-fold).In particular,the titer of MK-7 in a 50-L bioreactor reached 1549.6 mg·L^(-1),representing the highest titer reported so far.Thus,we propose a TIMO genetic circuits-assisted IPP metabolism remodeling framework that can be generally used for the synergistic fine-tuning of complicated metabolic modules to achieve the efficient bioproduction of terpenoids.

航空发动机用树脂基复合材料应用进展与发展趋势

树脂基复合材料具有比强度和比模量高、疲劳性能和耐腐蚀性能好等优点,已经成为航空发动机冷端部件的应用和发展趋势。国外航空发动机用树脂基复合材料研究起步较早,已经在多型发动机的风扇叶片、风扇机匣、外涵机匣、短舱等部件得到成熟应用,并朝着结构形式更优、材料性能更好、制造成本更低、自动化程度更高的方向发展。国内树脂基复合材料发展基础良好,但与国外相比在发动机上应用比例不高,需要进一步提升设计、材料、制造、实验技术水平及工程化能力。本文重点论述国外航空发动机复合材料构件的结构、材料和工艺发展现状,分析发展趋势,从建立航空发动机用复合材料体系、加强应用研究和设计牵引、推进预研成果转化和自动化技术应用等方面提出相关建议。

Food synthetic biology-driven protein supply transition: From animal-derived production to microbial fermentation

Animal-derived protein production is one of the major traditional protein supply methods,which continues to face increasing challenges to satisfy global needs due to population growth,augmented individual protein consumption,and aggravated environmental pollution.Thus,ensuring a sustainable protein source is a considerable challenge.The emergence and development of food synthetic biology has enabled the establishment of cell factories that effectively synthesize proteins,which is an important way to solve the protein supply problem.This review aims to discuss the existing problems of traditional protein supply and to elucidate the feasibility of synthetic biology in the process of protein synthesis.Moreover,using artificial bioengineered milk and artificial bioengineered eggs as examples,the progress of food protein supply transition based on synthetic biology has been systematically summarized.Additionally,the future of food synthetic biology as a potential source of protein has been also discussed.By strengthening and innovating the application of food synthetic biology technologies,including genetic engineering and high-throughput screening methods,the current limitations of artificial foods for protein synthesis and production should be addressed.Therefore,the development and industrial production of new food resources should be explored to ensure safe,high-quality,and sustainable global protein supply.

Nitrogen and Boron Co-Doped Carbon Nanotubes Embedded with Nickel Nanoparticles as Highly Efficient Electromagnetic Wave Absorbing Materials

Due to the limitations of impedance matching and attenuation matching,carbon nanotubes(CNTs)employed alone have a weak capacity to attenuate electromagnetic wave(EMW)energy.In this work,B and N co-doped CNTs with embedded Ni nanoparticles(Ni@BNCNTs)are fabricated via an in situ doping method.Compared with a sample without B doping,Ni@BNCNTs demonstrate a superior EMW absorption performance,with all minimum reflection loss values below−20 dB,even at a matching thickness of 1.5 mm.The experimental and theoretical calculation results demonstrate that B doping increases conduction and polarization relaxation losses,as well as the impedance matching characteristic,which is responsible for the enhanced EMW absorption performance of Ni@BNCNTs.

Henan Cultural Information Resources Sharing Project Persistent Effect Mechanism Research

In order to reduce east--west section area of “digital divider” in China, Ministry of Culture laun- ches the National Cultural Information Resources Sharing Project in the mid-west provinces, and the state finance gives certain funds subsidy in the project construction period. Embarks highly from strategy during construction plan chosen and operational system management, as while as union digital library to construct a persistent effect mechanism. With the principle of state guidance, enterprise operation and benefit farmers, expand culture industry within the provincial and safeguard Cultural Information Resources Sharing Project sustainable development's, then promotes the rural economy culture full scale development.

Effect and Safety of Iron Supplementation on Mild Thalassemia Complicated by Iron Deficiency in Late Pregnancy

Aim:To study the clinical effect and safety of iron supplementation in the treatment of mild thalassemia complicated by iron deficiency in late pregnancy.Methods:376 patients with mild thalassemia complicated by iron deficiency in late pregnancy treated in our hospital from July 2019 to June 2021 were selected and recruited in the research group,and 200 normal pregnant women treated in the same period were selected and recruited in the control group.Hemoglobin(Hb),reticulocyte blood,red blood protein(RET he)level,ferritin(SF)level,soluble transferrin receptor(sTfR)level,adverse maternal and infant outcomes,etc.were analyzed.Results:There was no significant difference in the levels of SF,Hb,RET he and sTfR between the two groups before treatment.After treatment,the levels of SF,Hb,RET he and sTfR in the research group were significantly improved,which was statistically significant compared with the control group.The frequencies of intrapartum hemorrhage,premature delivery,stillbirth,hemorrhagic shock,and neonatal asphyxia in the research group were 289±47.88,36(9.57%),0(0.00%),25(6.65%),and 6(1.26%),respectively.The frequencies of intrapartum hemorrhage,premature delivery,stillbirth,hemorrhagic shock,and neonatal asphyxia in the control group were 284±46.99,7(3.50%),0(0.00%),6(3.00%),and 0(0.00%),respectively.There were significant differences in preterm delivery and hemorrhagic shock,and there was no significant difference in other delivery outcomes.Conclusions:Under the premise of strictly controlling the iron reserve in pregnant women,continuous medication until the end of delivery may be conducive to the control of maternal thalassemia complicated by iron deficiency and the improvement of pregnancy outcome.

Efficient biosynthesis of creatine by whole-cell catalysis from guanidinoacetic acid in Corynebacterium glutamicum

Creatine is a naturally occurring derivative of an amino acid commonly utilized in functional foods and pharmaceuticals.Nevertheless,the current industrial synthesis of creatine relies on chemical processes,which may hinder its utilization in certain applications.Therefore,a biological approach was devised that employs whole-cell biocatalysis in the bacterium Corynebacterium glutamicum,which is considered safe for use in food production,to produce safe-for-consumption creatine.The objective of this study was to identify a guanidinoacetate N-methyltransferase(GAMT)with superior catalytic activity for creatine production.Through employing whole-cell biocatalysis,a gamt gene from Mus caroli(Mcgamt)was cloned and expressed in C.glutamicum ATCC 13032,resulting in a creatine titer of 3.37 g/L.Additionally,the study employed a promoter screening strategy that utilized nine native strong promoters in C.glutamicum to enhance the expression level of GAMT.The highest titer was achieved using the P1676 promoter,reaching 4.14 g/L.The conditions of whole-cell biocatalysis were further optimized,resulting in a creatine titer of 5.42 g/L.This is the first report of successful secretory creatine expression in C.glutamicum,which provides a safer and eco-friendly approach for the industrial production of creatine.

Facilitating stable gene integration expression and copy number amplification in Bacillus subtilis through a reversible homologous recombination switch

Strengthening the expression level of integrated genes on the genome is crucial for consistently expressing key enzymes in microbial cell factories for efficient bioproduction in synthetic biology.In comparison to plasmid-based multi-copy expression,the utilization of chromosomal multi-copy genes offers increased stability of expression level,diminishes the metabolic burden on host cells,and enhances overall genetic stability.In this study,we developed the“BacAmp”,a stabilized gene integration expression and copy number amplification system for high-level expression in Bacillus subtilis,which was achieved by employing a combination of repressor and non-natural amino acids(ncAA)-dependent expression system to create a reversible switch to control the key gene recA for homologous recombination.When the reversible switch is turned on,genome editing and gene amplification can be achieved.Subsequently,the reversible switch was turned off therefore stabilizing the gene copy number.The stabilized gene amplification system marked by green fluorescent protein,achieved a 3-fold increase in gene expression by gene amplification and maintained the average gene copy number at 10 after 110 generations.When we implemented the gene amplification system for the regulation of N-acetylneuraminic acid(NeuAc)synthesis,the copy number of the critical gene increased to an average of 7.7,which yielded a 1.3-fold NeuAc titer.Our research provides a new avenue for gene expression in synthetic biology and can be applied in metabolic engineering in B.subtilis.

Expression and antimicrobial activity of the recombinant bovine lactoferricin in Pichia pastoris

Lactoferricin,a multifunctional peptide located in the N-terminal region of lactoferrin,has a broad-spectrum bacteriostatic activity.It is a promising candidate as a food additive and immune fortification agent and does not have the risks associated with drug residues and drug resistance.First,we performed promoter and host cell screening to achieve the recombinant expression of lactoferricin in Pichia pastoris,showing an initial titer of 19.5 mg/L in P.pastoris X-33 using PAOX1 promoter.Second,we constructed a 0030-α hybrid signal peptide by fusing the 0030 signal peptide with the pro-sequence of α-factor secretory signal peptide.This further increased the production of lactoferricin,with a titer of 28.8 mg/L in the fermentation supernatant in the shaking flask.Next,we increased the expression of lactoferricin by fusing it with anionic antioxidant peptides.The neutralization of positive charges yielded a titer of 55.3 mg/L in the shaking flask,and a highest titer of 193.9 mg/L in a 3-L bioreactor.The antimicrobial activity analysis showed that recombinant-expressed lactoferricin exhibited potent antibacterial activity against Escherichia coli,Bacillus subtilis,and Staphylococcus aureus.This study provides a reference for the construction of microbial cell factories capable of efficiently synthesizing antimicrobial peptides.

Author correction to“Immune-awakening Saccharomyces-inspired nanocarrier for oral target delivery to lymph and tumors”[Acta Pharmaceutica Sinica B 12(2022)4501-4518]

The authors regret that there is an error in the article Fig.9D1 due to the mistake of copying and pasting in the process of assembling figures and negligence in the proofreading,and also a mislabled error in the supporting Information Fig.S9 and S10.

Spatiotemporal Variability in Extreme Temperature Events in an Arid-Semiarid Region of China and Their Teleconnections with Large-Scale Atmospheric Circulation

With a warming climate,temperature extremes have been a main global issue in recent decades due to their potential influence on the sustainable development of human life and natural ecosystems.In this study,12 indicators of extreme temperature events are used to evaluate the spatiotemporal distribution,periodic structure and teleconnections with large-scale atmospheric circulation in Xinjiang,Northwest China by combining wavelet coherence(WTC) analysis based on continuous wavelet transform(CWT) analysis with the sequential Mann-Kendall test.We find that over the past six decades,the climate in Xinjiang has become warmer and has suffered from increases in the frequency of warm extremes and decreases in the frequency of cold extremes.Warm extremes have mainly occurred in the southern Tianshan Mountains surrounding the Tarim Basin and western part of the Taklamakan Desert,and cold extremes have primarily occurred in the southwestern Altai Mountains and northern foot of the Tianshan Mountains.Extreme temperature events,including warm extremes,cold extremes,and other temperature indices,have significant interannual variability,with the main oscillation periods at smaller(2–4-year band),intermediate(4–7-year band),and greater time scales in recent decades.Furthermore,cold-extreme indices,including frost days,cool days,and cool nights all show a clear changepoint during 1990–1997 at the 95% confidence level,and both ice days and cold spell duration indicator have a potential changepoint during 1981–1986.However,the changing points for warmextreme indices are detected during 1992–1998.The temperature variables are significantly correlated with the EI Ni?o-Southern Oscillation(ENSO) and Arctic Oscillation(AO),but less well correlated with the Pacific Decadal Oscillation(PDO).The phase difference in the WTC spectra is not uniform between temperature extremes and climatic oscillations.Our findings will have important implications for local governments in taking effective measures to mitigate the potential effects of regional climate warming due to human activities in Xinjiang.

Non-uniform operative temperature distribution characteristics and heat-source-controlled core-area range of local heating radiators

Heating the whole space,which is currently used in northern China,leads to high energy consumption and substantial pollution.A transition to local heating has the potential to help address this problem.In this paper,the effects of radiator-related parameters(position,power,and size)and room-related parameters(aspect ratio and height)on local heating were studied.Two evaluation indices,the effective coefficient of operative temperature(OTEC)and the effective coefficient of local heating(LHEC),were proposed.In addition,the heat source-control core-area(HSCCA)was proposed,and the effect range of heat sources in the space was evaluated by the attenuation of operative temperature.The findings demonstrated that the radiator position has a greater influence on local heating than size.When the position of the radiator was changed from"close to the inner wall"to"close to the outer wall",the LHEC(the interior one-quarter of room is a local heating zone)was found to decrease by 73%.The size of the radiator,which is close to the inner wall,doubled or quadrupled,and the LHEC increased by 9%and 18%.Moreover,rooms with a larger aspect ratio or small room height were found to be the most optimal for local heating applications.The area of the HSCCA decreased as the position of the radiator approached the outer wall.The findings of this study can be used as a design reference for the radiator when the heating mode changes from"full-space heating"to"local heating".

In silico cell factory design driven by comprehensive genome‑scale metabolic models:development and challenges

Genome-scale metabolic models(GEMs)have been widely used to design cell factories in silico.However,initial flux balance analysis only considers stoichiometry and reaction direction constraints,so it cannot accurately describe the distribution of metabolic flux under the control of various regulatory mechanisms.In the recent years,by introducing enzymology,thermodynamics,and other multiomics-based constraints into GEMs,the metabolic state of cells under different conditions was more accurately simulated and a series of algorithms have been presented for microbial phenotypic analysis.Herein,the development of multiconstrained GEMs was reviewed by taking the constraints of enzyme kinetics,thermodynamics,and transcriptional regulatory mechanisms as examples.This review focused on introducing and summarizing GEMs application tools and cases in cell factory design.The challenges and prospects of GEMs development were also discussed.

Experimental study on the influence of temperature and humidity on the thermal conductivity of building insulation materials

At present,thermal conductivity is usually taken as a constant value in the calculation of building energy con-sumption and load.However,in the actual use of building materials,they are exposed to the environment with continuously changing temperature and relative humidity.The thermal conductivity of materials will inevitably change with temperature and humidity,leading to deviations in the estimation of energy consumption in the building.Therefore,in this study,variations in the thermal conductivity of eight common building insulation materials(glass wool,rock wool,silica aerogel blanket,expanded polystyrene,extruded polystyrene,phenolic foam,foam ceramic and foam glass)with temperature(in the range of 20-60°C)and relative humidity(in the range of 0-100%)were studied by experimental methods.The results show that the thermal conductivity of these common building insulation materials increased approximately linearly with increasing temperature with maxi-mum growth rates from 3.9 to 22.7%in the examined temperature range.Due to the structural characteristics of materials,the increasing thermal conductivity of different materials varies depending on the relative humidity.The maximum growth rates of thermal conductivity with humidity ranged from 8.2 to 186.7%.In addition,the principles of selection of building insulation materials in different humidity regions were given.The research re-sults of this paper aim to provide basic data for the accurate value of thermal conductivity of building insulation materials and for the calculation of energy consumption.

Improving the Estimation of Salt Distribution during Evaporation in Saline Soil by HP1 Model

Restricted by the development of the transient flow and solute reactive transport models for unsaturated soil, empirical functions have been used previously to calculate the mass of dissolved or precipitated salt when they have to be taken into account. Besides, the solute reactive transport process has often been inferred based on measurements that cost lots of time and manpower. HP1 model coupled with PHREEQC provides a suitable tool to improve the estimation of salt distribution during evaporation in saline soil, where the salt dissolution and precipitation cannot be ignored. In this study, we compare the performance of a standard solute transport(SST) model and the HP1 model to examine the improvement of salt distribution estimation. Model results are compared with experimental data sets from four field lysimeters. These columns were exposed to Na Cl solution with different concentrations(3, 30, 100, and 250 g/L) and were undergoing the same strong evaporation boundary condition. The pre-existing Ca SO_(4), Na Cl and Na2SO_(4)loads were 1.15, 0.47 and 0.23 g/(100 g of soil), respectively. Simulation results show that HP1 ameliorates the overestimation of salt content by SST in deeper soil due to the absence of dissolution of pre-existing soluble salts, and prevents the concentration of the solute from exceeding the solubilities which would occur in SST-result. Additionally, HP1-predicted results can help trace the transport process of each solute. Based on the results, we strongly suggest that the management of fields sensitive to salt content should make use of a coupled flow and chemical reaction model.

Thermal performance and evaluation of a novel stratified and mixed flexible transformation solar heat storage unit

It is necessary to satisfy the flexible requirements of solar heat storage systems to provide efficient heating and constant-temperature domestic hot water at different periods.A novel heat storage tank with both stratified and mixing functions is proposed,which can realize the integration of stable stratification and rapid mixing modes.In this research,a three-dimensional heat transfer model of the heat storage tank with stratified and mixed dual modes was established,and a thermal performance test system for the tank was built in the State Key Laboratory of Green Building in Western China.Moreover,a new evaluation index representing the mixing speed is proposed.The stratification effect and mixing characteristics of the tank were studied under different comprehensive conditions.The results show that the exergy efficiency of the tank with a stratified pipe can be increased by 10%–15%compared to that of a conventional tank.Additionally,the recommended optimal flow rate range for well-stratified tanks is 4–6 L/min.The mixing nozzle of the tank reduces the mixing reaction coefficient by 0.27 and significantly reduces the mixing time.This study provides critical guidance to meet the flexible thermal needs of users and implement high-performance applications using the stratified and mixing modes of heat storage tanks.

Timetabling optimization of classrooms and self-study rooms in university teaching buildings based on the building controls virtual test bed platform considering energy efficiency

The energy consumption of a teaching building can be effectively reduced by timetable optimization.However,in most studies that explore methods to reduce building energy consumption by course timetable optimization,self-study activities are not considered.In this study,an MATLAB-EnergyPlus joint simulation model was constructed based on the Building Controls Virtual Test Bed platform to reduce building energy consumption by optimizing the course schedule and opening strategy of self-study rooms in a holistic way.The following results were obtained by taking a university in Xi’an as an example:(1)The energy saving percentages obtained by timetabling optimization during the heating season examination week,heating season non-examination week,cooling season examination week,and cooling season non-examination week are 35%,29.4%,13.4%,and 13.4%,respectively.(2)Regarding the temporal arrangement,most courses are scheduled in the morning during the cooling season and afternoon during the heating season.Regarding the spatial arrangement,most courses are arranged in the central section of the middle floors of the building.(3)During the heating season,the additional building energy consumption incurred by the opening of self-study rooms decreases when duty heating temperature increases.

Multi-objective optimization of equipment capacity and heating network design for a centralized solar district heating system

Northwest China has abundant solar energy resources and a large demand for winter heating.Using solar energy for centralized heating is a clean and effective way to solve local heating problems.While present studies usually decoupled solar heating stations and the heating network in the optimization design of centralized solar heating systems,this study developed a joint multi-objective optimization model for the equipment capacity and the diameters of the heating network pipes of a centralized solar district heating system,using minimum total life cycle cost and CO_(2)emission of the system as the optimization objectives.Three typical cities in northwest China with different solar resource conditions(Lhasa,Xining,and Xi'an)were selected as cases for analysis.According to the results,the solar heating system designed using the method proposed in this study presents lower economic cost and higher environmental protection in comparison to separately optimizing the design of the solar heating station and the heating network.Furthermore,the solar fraction of the optimal systems are 90%,70%,and 31%for Lhasa,Xining,and Xi'an,and the minimum water supply temperatures are 55℃,50℃,and 65℃for an optimal economy and 55℃,45℃,and 45℃for optimal environmental protection,respectively.It was also established that the solar collector price has a greater impact on the equipment capacity of the solar heating station than the gas boiler price.

Crude enzyme immobilization‑based cell‑free system for efficient N‑acetylneuraminic acid biosynthesis aided by N‑terminal coding sequence screening

N-acetylneuraminic acid(NeuAc)is an important nutrient that plays a key role in brain development in infants NeuAc is mainly produced by extraction from natural resources such as edible birds’s nests,crucian eggs,caviar and human breast milk.The extraction process is complicated,resulting in the disadvantages of low NeuAc content and low recovery rate.In this study,a crude enzyme immobilization-based cell-free system(CEICFS)was developed for efficient NeuAc biosynthesis.First,N-terminal coding sequences that improved the expression levels of N-acetylglucosamine-2-epimerase(AGE)and N-acetylneuraminic acid aldolase(NanA)were obtained by high-throughput screening.And these sequences resulted in up to 1.5-fold(1.2-fold)increase in AGE(NanA)enzyme levels.And then,a CEICFS for NeuAc biosynthesis was proposed by directly immobilizing crude enzyme containing AGE and NanA on amino resin.Subsequently,NeuAc production from GlcNAc using CEICFS in one reactor was carried out,resulting 68 g/L of NeuAc and the highest productivity of 6.8 g/L/h.Further,the enzyme activity was still higher than 75%after five repeated uses.The functional properties of CEICFS were studied and compared to those of the free enzyme,immobilization can extend the application of enzyme to some harsh environments,such as low temperature and acidic environment.Therefore,CEICFS with excellent heat resistance,storage stability and reusability exhibit great potential for industrial application.