Light-driven CO2 conversion system: construction, optimization and application / 生物工程学报

The use of light energy to drive carbon dioxide (CO2) reduction for production of chemicals is of great significance for relieving environmental pressure and solving energy crisis. Photocapture, photoelectricity conversion and CO2 fixation are the key factors affecting the efficiency of photosynthesis, and thus also affect the efficiency of CO2 utilization. To solve the above problems, this review systematically summarizes the construction, optimization and application of light-driven hybrid system from the perspective of combining biochemistry and metabolic engineering. We introduce the latest research progress of light-driven CO2 reduction for biosynthesis of chemicals from three aspects: enzyme hybrid system, biological hybrid system and application of these hybrid system. In the aspect of enzyme hybrid system, many strategies were adopted such as improving enzyme catalytic activity and enhancing enzyme stability. In the aspect of biological hybrid system, many methods were used including enhancing biological light harvesting capacity, optimizing reducing power supply and improving energy regeneration. In terms of the applications, hybrid systems have been used in the production of one-carbon compounds, biofuels and biofoods. Finally, the future development direction of artificial photosynthetic system is prospected from the aspects of nanomaterials (including organic and inorganic materials) and biocatalysts (including enzymes and microorganisms).

Metabolic engineering of Escherichia coli for adipic acid production / 生物工程学报

Adipic acid is a high-value-added dicarboxylic acid which is primarily used in the production of nylon-66 for manufacturing polyurethane foam and polyester resins. At present, the biosynthesis of adipic acid is hampered by its low production efficiency. By introducing the key enzymes of adipic acid reverse degradation pathway into a succinic acid overproducing strain Escherichia coli FMME N-2, an engineered E. coli JL00 capable of producing 0.34 g/L adipic acid was constructed. Subsequently, the expression level of the rate-limiting enzyme was optimized and the adipic acid titer in shake-flask fermentation increased to 0.87 g/L. Moreover, the supply of precursors was balanced by a combinatorial strategy consisting of deletion of sucD, over-expression of acs, and mutation of lpd, and the adipic acid titer of the resulting E. coli JL12 increased to 1.51 g/L. Finally, the fermentation process was optimized in a 5 L fermenter. After 72 h fed-batch fermentation, adipic acid titer reached 22.3 g/L with a yield of 0.25 g/g and a productivity of 0.31 g/(L·h). This work may serve as a technical reference for the biosynthesis of various dicarboxylic acids.

Modular engineering of Escherichia coli for high-level production of l-tryptophan / 生物工程学报

As an essential amino acid, l-tryptophan is widely used in food, feed and medicine sectors. Nowadays, microbial l-tryptophan production suffers from low productivity and yield. Here we construct a chassis E. coli TRP3 producing 11.80 g/L l-tryptophan, which was generated by knocking out the l-tryptophan operon repressor protein (trpR) and the l-tryptophan attenuator (trpL), and introducing the feedback-resistant mutant aroGfbr. On this basis, the l-tryptophan biosynthesis pathway was divided into three modules, including the central metabolic pathway module, the shikimic acid pathway to chorismate module and the chorismate to tryptophan module. Then we used promoter engineering approach to balance the three modules and obtained an engineered E. coli TRP9. After fed-batch cultures in a 5 L fermentor, tryptophan titer reached to 36.08 g/L, with a yield of 18.55%, which reached 81.7% of the maximum theoretical yield. The tryptophan producing strain with high yield laid a good foundation for large-scale production of tryptophan.

Research progress in the role of distal femoral valgus cut angle in accurate total knee arthroplasty / 中华创伤杂志

The distal femoral fracture, tibial plateau fracture and patellar fracture may all develop into traumatic knee arthritis, which is probably associated with knee dysfunction problems. Total knee arthroplasty (TKA) is an effective treatment for end-stage knee arthritis. The distal femoral valgus cut angle (VCA) is an important reference for distal femoral resection in TKA and significantly affects postoperative prosthesis position and lower extremity alignment after TKA. For VCA, the specific methods for definition, measurement methods, clinical application and influencing factors are currently controversial. Hence, the authors review the research progress in the role of VCA in TKA from the above-mentioned four aspects, hoping to provide a reference for accurate preoperative planning and intraoperative performance of TKA.

Application and research progress of intelligent accuracy technological aids in total knee arthroplasty / 中华创伤杂志

Total knee arthroplasty (TKA) is an effective treatment for end-stage knee disease, with the postoperative alignment, component position, soft tissue balance, and prosthesis matching being key factors for the success of TKA. In order to achieve more accurate postoperative alignment and component position, better soft tissue balance and prosthesis matching for longer prosthesis longevity, better postoperative function and higher patient satisfaction, various intelligent accuracy technological aids such as computer assisted navigation (CAN), patient specific instrumentation (PSI), surgical robots, microsensors, customized implants (CI) and personalized 3D preoperative planning have emerged and are given high expectation. In this paper, the authors review the application and research progress of the above technological aids mainly from aspects of alignment, component position, clinical outcomes and cost analysis, so as to provide a reference for the application of related technological aids in TKA.

Advances in synthetic biology of CO2 fixation by heterotrophic microorganisms / 生物工程学报

Human activities increase the concentration of atmospheric carbon dioxide (CO2), which leads to global climate warming. Microbial CO2 fixation is a promising green approach for carbon neutral. In contrast to autotrophic microorganisms, heterotrophic microorganisms are characterized by fast growth and ease of genetic modification, but the efficiency of CO2 fixation is still limited. In the past decade, synthetic biology-based enhancement of heterotrophic CO2 fixation has drawn wide attention, including the optimization of energy supply, modification of carboxylation pathway, and heterotrophic microorganisms-based indirect CO2 fixation. This review focuses on the research progress in CO2 fixation by heterotrophic microorganisms, which is expected to serve as a reference for peaking CO2 emission and achieving carbon neutral by microbial CO2 fixation.

Association of 24-hour urinary sodium excretion and 25-hydroxy vitamin D level with albuminuria in patients with type 2 diabetes / 中华内分泌代谢杂志

Objective:Using 24-hour urinary sodium excretion (24h-UNa) as the surrogate measure of sodium intake, to evaluate the joint association of 24h-UNa and serum 25-hydroxy vitamin D (25-OHD) levels with the risk of albuminuria in patients with type 2 diabetes mellitus (T2DM).Methods:This retrospective study included 670 hospitalized T2DM cases in the Department of Endocrinology and Metabolic Diseases, the First Affiliated Hospital of Zhengzhou University from January 2018 to October 2021. Patients were divided into the albuminuria-positive group or negative-group according to the level of 24-hour urinary albumin excretion (24h-UAE); They were also divided into the high-sodium group or low-sodium group according to the level of 24h-UNa; Patients were divided into the low-VD group or high-VD group according to the level of 25-OHD. Combining 24h-UNa and 25-OHD, the patients were further divided into four groups: high-VD low-sodium group ( n=85), high-VD high-sodium group ( n=122), low-VD low-sodium group ( n=248), and low-VD high-sodium group ( n=215). The effect of 24h-UNa and 25-OHD association on albuminuria was analyzed by binary regression. Results:There were significant differences in 24h-UAE level among the four groups ( P<0.01), the level of 24h-UAE in the low-VD high-sodium group was significantly higher than that in low-VD low-sodium group, high-VD low-sodium group, and high-VD high-sodium group [39.00(13.00, 319.00)mg/24 h vs 22.00(10.00, 99.00)mg/24 h, 22.00(9.00, 72.50)mg/24 h, 22.45(9.69, 72.75)mg/24 h; P=0.047, P=0.019, P=0.030]. Correlation analysis showed a positive correlation between 24h-UNa and 24h-UAE in the low-VD group ( P=0.017), but not in the high-VD group ( P=0.411). Binary regression analyses showed that both 24h-UNa ( P=0.017) and 25-OHD( P=0.023) were independent risk factors for positive albuminuria in patients with T2DM. The risk of positive albuminuria in the low-VD high-sodium group was 1.789 times higher than that in the high-VD low-sodium group ( P=0.037). Conclusion:24h-UAE in T2DM patients was affected by the combination of 24h-UNa and 25-OHD. A low level of 25-OHD increased the risk of albuminuria in high sodium intake T2DM patients.

Application of chronological lifespan in the construction of Escherichia coli cell factories / 生物工程学报

Microbial cell factories capable of producing valuable chemicals from renewable feedstocks provide a promising alternative towards sustainability. However, environmental stress remarkably affects the performance of microbial cell factories. By extending the chronological lifespan of microbial cells, the performance of microbial cell factories can be greatly improved. Firstly, an evaluation system for chronological lifespan and semi-chronological lifespan was established based on the changes in survival rates. Secondly, the addition of anti-aging drugs such as cysteine, carnosine, aminoguanidine and glucosamine increased the chronological lifespan of E. coli by 80%, 80%, 50% and 120%, respectively. Finally, we demonstrated that extending the chronological lifespan of E. coli increased the yield of metabolites produced by E. coli cell factories with endogenous (lactic acid and pyruvic acid) or exogenous (malic acid) metabolic pathway by 30.0%, 25.0%, and 27.0%, respectively. The strategy of extending chronological lifespan of E. coli provides a potential approach for enhancing the performance of microbial cell factories.

Biosynthesis of 2,5-dimethylpyrazine from L-threonine by whole-cell biocatalyst of recombinant Escherichia coli / 生物工程学报

2,5-dimethylpyrazine (2,5-DMP) is of important economic value in food industry and pharmaceutical industry, and is now commonly produced by chemical synthesis. In this study, a recombinant Escherichia coli high-efficiently converting L-threonine to 2,5-DMP was constructed by combination of metabolic engineering and cofactor engineering. To do this, the effect of different threonine dehydrogenase (TDH) on 2,5-DMP production was investigated, and the results indicate that overexpression of EcTDH in E. coli BL21(DE3) was beneficial to construct a 2,5-DMP producer with highest 2,5-DMP production. The recombinant strain E. coli pRSFDuet-tdh(Ec) produced (438.3±23.7) mg/L of 2,5-DMP. Furthermore, the expression mode of NADH oxidase (NoxE) from Lactococcus cremoris was optimized, and fusion expression of EcTDH and LcNoxE led to balance the intracellular NADH/NAD⁺ level and to maintain the high survival rate of cells, thus further increasing 2,5-DMP production. Finally, the accumulation of by-products was significantly decreased because of disruption of shunt metabolic pathway, thereby increasing 2,5-DMP production and the conversion ratio of L-threonine. Combination of these genetic modifications resulted in an engineered E. coli Δkbl ΔtynA ΔtdcB ΔilvA pRSFDuet-tdhEcnoxELc-PsstT (EcΔkΔAΔBΔA/TDH(Ec)NoxE(Lc)-PSstT) capable of producing (1 095.7±81.3) mg/L 2,5-DMP with conversion ratio of L-threonine of 76% and a yield of 2,5-DMP of 28.8% in 50 mL transformation system with 5 g/L L-threonine at 37 °C and 200 r/min for 24 h. Therefore, this study provides a recombinant E. coli with high-efficiently catalyzing L-threonine to biosynthesize 2,5-DMP, which can be potentially used in biosynthesis of 2,5-DMP in industry.

Early and mid-term outcomes of concomitant cryosurgical Cox-Maze Ⅳ procedure in minimally invasive mitral valve surgery / 中国胸心血管外科临床杂志

@#Objective    To analyze the early and mid-term safety and effectiveness of concomitant cryosurgical Cox-Maze Ⅳ procedure in minimally invasive mitral valve surgery. Methods    We retrospectively reviewed the clinical data of 68 patients (28 males and 40 females with a mean age of 38.7±9.3 years) who underwent concomitant cryosurgical Cox-Maze Ⅳ procedure in minimally invasive mitral valve and tricuspid surgery in the Department of Cardiovascular Surgery of the Second Xiangya Hospital from August 2013 to October 2017. The heart rhythm of the patients after surgery was supervised by 24 hour holter monitoring eletrocardiogram. Results    No death occurred during operation and follow-up. One patient underwent reexploration for bleeding. The rate of sinus rhythm restored at the time of discharge was 95.8%. The rate of sinus rhythm restored at 6 months, 12 months, 24 months, 36 months after surgery was 93.5%, 91.6%, 90.3% and 89.5% respectively. Conclusion    Concomitant cryosurgical Cox-Maze Ⅳ procedure in minimally invasive mitral valve surgery is quite safe and effective in treatment of rheumatic mitral valve disease and atrial fibrillation in the early and mid-term follow-up.

The data of Chinese minimally invasive cardiovascular surgery in 2019 / 中国胸心血管外科临床杂志

@#The minimally invasive cardiovascular surgery developed rapidly in last decades. In order to promote the development of minimally invasive cardiovascular surgery in China, the Chinese Minimally Invasive Cardiovascular Surgery Committee (CMICS) has gradually standardized the collection and report of the data of Chinese minimally invasive cardiovascular surgery since its establishment. The total operation volume of minimally invasive cardiovascular surgery in China has achieved substantial growth with a remarkable popularization of concepts of minimally invasive medicine in 2019. The data of Chinese minimally invasive cardiovascular surgery in 2019 was reported as a paper for the first time, which may provide reference to cardiovascular surgeons and related professionals.

Production of L-2-aminobutyric acid from L-threonine using a trienzyme cascade / 生物工程学报

L-2-aminobutyric acid (L-ABA) is an important chemical raw material and chiral pharmaceutical intermediate. The aim of this study was to develop an efficient method for L-ABA production from L-threonine using a trienzyme cascade route with Threonine deaminase (TD) from Escherichia. coli, Leucine dehydrogenase (LDH) from Bacillus thuringiensis and Formate dehydrogenase (FDH) from Candida boidinii. In order to simplify the production process, the activity ratio of TD, LDH and FDH was 1:1:0.2 after combining different activity ratios in the system in vitro. The above ratio was achieved in the recombinant strain E. coli 3FT+L. Moreover, the transformation conditions were optimized. Finally, we achieved L-ABA production of 68.5 g/L with a conversion rate of 99.0% for 12 h in a 30-L bioreactor by whole-cell catalyst. The environmentally safe and efficient process route represents a promising strategy for large-scale L-ABA production in the future.

Using dynamic molecular switches for shikimic acid production in Escherichia coli / 生物工程学报

Shikimic acid is an intermediate metabolite in the synthesis of aromatic amino acids in Escherichia coli and a synthetic precursor of Tamiflu. The biosynthesis of shikimic acid requires blocking the downstream shikimic acid consuming pathway that leads to inefficient production and cell growth inhibition. In this study, a dynamic molecular switch was constructed by using growth phase-dependent promoters and degrons. This dynamic molecular switch was used to uncouple cell growth from shikimic acid synthesis, resulting in the production of 14.33 g/L shikimic acid after 72 h fermentation. These results show that the dynamic molecular switch could redirect the carbon flux by regulating the abundance of target enzymes, for better production.

Recent advances in enzymatic production of alpha-keto acids / 生物工程学报

Alpha-keto acid is a bifunctional organic compound containing both carboxyl and ketone groups, and widely applied in the industries of food, pharmaceutical and cosmetics. Based on the demand of eco-friendly process, safety and sustainable development, production of α-keto acids by enzymatic conversion technology has been paid more and more attention. In this article, we review the status of α-keto acids biosynthesis from three aspects: enzymatic screening, enzymatic modification and optimization of enzymatic conversion conditions. Meanwhile, we also indicate future research directions for further improving α-keto acids production.

Application of metabolic network model to analyze intracellular metabolism of industrial microorganisms / 生物工程学报

To quickly and efficiently understand the intracellular metabolic characteristics of industrial microorganisms, and to find potential metabolic engineering targets, genome-scale metabolic network models (GSMMs) as a systems biology tool, are attracting more and more attention. We review here the 20-year history of metabolic network model, analyze the research status and development of GSMMs, summarize the methods for model construction and analysis, and emphasize the applications of metabolic network model for analyzing intracellular metabolic activity of microorganisms from cellular phenotypes, and metabolic engineering. Furthermore, we indicate future development trend of metabolic network model.

Development of a purification tag to produce thermostable fused protein / 生物工程学报

Self-assembling amphipathic peptides (SAPs) have alternating hydrophilic and hydrophobic residues and can affect the thermal stabilities and catalytic properties of the fused enzymes. In this study, a novel multifunctional tag, S1vw (HNANARARHNANARARHNANARARHNARARAR) was developed to modify fused enzymes. After fusing S1vw at the enzymes/proteins N-terminus through a PT-linker, the crude enzymatic activities of polygalacturonate lyase and lipoxygenase were enhanced 3.1- and 1.89-fold, respectively, compared to the wild-type proteins. The relative fluorescence intensity of the green fluorescent protein was enhanced 16.22-fold. All the three S1vw fusions could be purified by nickel column with high purities and acceptable recovery rates. Moreover, S1vw also induced the thermostabilities enhancement of the fusions, with polygalacturonate lyase and lipoxygenase fusions exhibiting 2.16- and 3.2-fold increase compared with the corresponding wild-type, respectively. In addition, S1vw could enhance the production yield of green fluorescent protein in Escherichia coli and Bacillus subtilis while the production of GFP and its S1vw fusion changed slightly in Pichia pastoris. These results indicated that S1vw could be used as a multifunctional tag to benefit the production, thermal stability and purification of the fusion protein in prokaryotic expression system.

Optimization of Preparation Technology of Near-infrared Light Response-based Doxorubicin Hydrochloride Nano-liposomes / 中国药房

OBJECTIVE： To establish a methaod for content determination of doxorubicin hydrochloride nano-liposomes， and to optimize its preparation technology. METHODS： The contents of doxorubicin hydrochloride nano-liposomes was determined by UV spectrophotometry. The membrane dispersion method was used to prepare doxorubicin hydrochloride nano-liposomes. Using particle size， encapsulation efficiency and drug-loading amount as indexes， the weight ratio of phospholipid to drug （mg/mg）， the weight ratio of phospholipid to cholesterol （mg/mg） and ultrasonic time （min） as factors， central composite design-response surface methodology was used to optimize the preparation technology. The photothermal conversion effect of doxorubicin hydrochloride nano-liposomes was investigated by near infrared irradiation. RESULTS： The linear range of doxorubicin hydrochloride were 1.01-16.16 μg/mL（r＝0.999 7）； precision， stability and reproducibility tests were all in line with the requirments of Chinese Pharmacopoeia. The optimal preparation technology included that the weight ratio of phospholipid to drug was 13.30 ∶ 1（mg/mg）； the weight ratio of phospholipid to cholesterol was 4.09 ∶ 1 （mg/mg）； the ultrasonic time was 10 min. Under this technology， the particle size and drug-loading amount of doxorubicin hydrochloride nano-liposomes were （200.5±25.1） nm and （11.02±0.20）％， relative errors of which to predicted value （196.3 nm， 10.68％） were 1.82％ and 1.63％. The consistency between measured value and predicted value was good. Doxorubicin hydrochloride nano-liposomes exhibited concentration- dependent and time-dependent photothermal conversion characteristics under near infrared irradiation at 808 nm. CONCLUSIONS： Established method is simple and good accuracy. The optimized preparation technology is simple and feasible.

High frequency repeated transcranial magnetic stimulation improves learning and memory of global cerebral ischemia rats and its mechanism / 中华神经科杂志

Objective To study the effect of high frequency repeated transcranial magnetic stimulation (rTMS) on learning and memory in rats with global cerebral ischemia and to investigate its mechanism.Methods Thirty-five male Sprague-Dawley rats (8-10 weeks old) were randomly divided into sham operation group (n=8),model group (n=9),sham-rTMS (s-rTMS) group (n=9) and rTMS group (n=9).The global cerebral ischemia model was established by modified four-vessel occlusion method.The rTMS group received 10 Hz rTMS stimulation for two weeks,whereas the s-rTMS group received sham stimulation.Morris water maze test was used to detect the spatial learning ability,multi-channel recording technique was used to detect the local field potentials in the hippocampus CA1 region of theta and gamma oscillation,and immunohistochemical staining and Western blotting were used to detect the expression of protein kinase A (PKA) and phosphorylated cyclic adenosine monophosphate response element binding protein (p-CREB) of hippocampus.Results The average escape latency in the model group was longer than that in the sham operation group ((35.16±0.80) s vs (16.57±0.74) s,k=3.723,P=0.013),the spanning platform times and the original platform quadrant swimming time in the model group were shorter than that in the sham operation group (1.14±0.42 vs 4.46±0.23,k=3.185,P=0.042;(14.46±0.73) s vs (29.31±0.42) s,k=3.027,P=0.047).Compared with the sham operation group,the mean power spectral density of theta and gamma reduced ((-68.48±2.61) Hz vs (-59.38±2.25) Hz,k=2.958,P=0.048;(-82.23±4.60) Hz vs (-70.50±4.25) Hz,k=3.729,P=0.021),and the expression of PKA and p-CREB protein decreased in the model group (7 184.26±975.12 vs 25 137.35±1 010.62,k=3.588,P=0.027;1 803.73±336.18 vs 20 175.25±727.23,k=2.912,P=0.049).The average escape latency in the rTMS group was shorter than that in the model group ((24.69± 1.01) s vs (35.16±0.80) s,k=4.082,P=0.034),and the spanning platform times and the original platform quadrant swimming time in the rTMS group was longer than that in the model group (2.42±0.31 vs 1.14±0.42,k=3.296,P=0.039;(23.07±0.67) s vs (14.46±0.73) s,k=4.323,P=0.012).Compared with the rTMS group,the power spectral density of theta and gamma reduced ((-63.81±3.12) Hz vs (-68.48±2.61) Hz,k=3.582,P=0.015;(-75.80±4.58) Hz vs (-82.23±4.60) Hz,k=4.051,P=0.026),and the expression of PKA and p-CREB protein decreased in the model group (13 065.32±1 045.18 vs 7 184.26±975.12,k=3.923,P=0.031;11 032.83±562.86 vs 18 03.73±336.18,k=3.178,P=0.038).Conclusion High frequency rTMS could improve learning and memory of global cerebral ischemia rats,the mechanism of which might be that rTMS enhance the hippocampal theta and gamma oscillations and increase the expression of PKA and p-CREB protein in the hippocampus,thus increasing the hippocampus synaptic plasticity.

Mutation and fermentation optimization of Bacillus amyloliquefaciens for acetoin production / 生物工程学报

As a platform chemical, acetoin has a great potential of application in medicine and food industries. In order to improve the efficiency of acetoin production, Bacillus amyloliquefaciens was treated by atmospheric and room temperature plasma and gamma rays. Two-round screening was adopted for obtaining positive mutants, and the best mutant B. amyloliquefaciens H-5 produced acetoin up to 68.2 g/L in shake flask. Then, culture conditions were optimized in 5-L fermentor to enhance acetoin production. Finally, 85.2 g/L acetoin was produced by B. amyloliquefaciens H-5, which was increased by 26.8% compared with that of the original strain B. amyloliquefaciens FMME088. These results indicated that the high-producing strain can be obtained efficiently by compound mutagenesis, which has a promising prospect for commercial scale process.

Pharmacokinetics of monoclonal antibodies and Fc-fusion proteins

There are many factors that can influence the pharmacokinetics (PK) of a mAb or Fc-fusion molecule with the primary determinant being FcRn-mediated recycling. Through Fab or Fc engineering, IgG-FcRn interaction can be used to generate a variety of therapeutic antibodies with significantly enhanced half-life or ability to remove unwanted antigen from circulation. Glycosylation of a mAb or Fc-fusion protein can have a significant impact on the PK of these molecules. mAb charge can be important and variants with pI values of 1-2 unit difference are likely to impact PK with lower pI values being favorable for a longer half-life. Most mAbs display target mediated drug disposition (TMDD), which can have significant consequences on the study designs of preclinical and clinical studies. The PK of mAb can also be influenced by anti-drug antibody (ADA) response and off-target binding, which require careful consideration during the discovery stage. mAbs are primarily absorbed through the lymphatics via convection and can be conveniently administered by the subcutaneous (sc) route in large doses/volumes with co-formulation of hyaluronidase. The human PK of a mAb can be reasonably estimated using cynomolgus monkey data and allometric scaling methods.