L-valine is an important branched-chain amino acid widely used in the food, pharmaceutical, and feed industries. Microbial fermentation has become the primary production method for L-valine. However, current industrial production still faces issues such as inefficient carbon flux utilization, imbalance in cofactor supply and demand, and suboptimal fermentation processes, which limit the efficient synthesis of L-valine. To further enhance the production performance of L-valine, In this study, metabolic engineering was conducted for a previously constructed Escherichia coli strain with a high yield of L-valine to optimize carbon flux distribution and balance cofactor consumption. Dual-phase oxygen-controlled fermentation was carried out to enhance L-valine production. Firstly, to address the pyruvate loss, we knocked out multiple competing pathway genes (ldhA, poxB, pflB, frdA, and pta), which resulted in a 48% increase in flask yield of the constructed strain VL-04. Next, we optimized the cofactor supply and demand balance by replacing ilvE with bcd (NADH-preferential) from Bacillus subtilis to construct the strain VL-06, which achieved a flask yield of 22.80 g/L, a further improvement of 25.8%. Subsequently, the fermentation conditions of VL-06 were optimized in a 5 L bioreactor with dual-phase oxygen-controlled fermentation. After optimization, the L-valine production reached 86.44 g/L in 26 h, with a glucose-to-acid conversion rate of 44.08% and a production intensity of 3.32 g/(L·h). This study not only shortens the time for L-valine production but also improves the economic efficiency, providing insights for similar fermentation processes employing dual-phase oxygen control.
Metabolic Engineering/methods* ; Escherichia coli/genetics* ; Valine/biosynthesis* ; Fermentation ; Bacillus subtilis/genetics*

Phenylpyruvic acid (PPA) is used as a food and feed additive and has a wide range of applications in the pharmaceutical, chemical and other fields. At present, PPA is mainly produced by chemical synthesis. With the green transformation of the manufacturing industry, biotransformation will be a good alternative for PPA production. The L-amino acid deaminase (PmiLAAD) from Proteus mirabilis has been widely studied for the production of PPA. However, the low yield limits its industrial production. To further enhance the production of PPA and better meet industrial demands, a more efficient synthesis method for PPA was established. In this study, PmiLAAD was heterologously expressed in Escherichia coli. Subsequently, a colorimetric reaction method was established to screen the strains with high PPA production. The semi-rational design of PmiLAAD was carried out, and the obtained triple-site mutant V18 (V437I/S93C/E417A) showed a 35% increase in catalytic activity compared with the wild type. Meanwhile, the effect of N-terminal truncation on the catalytic activity of the V18 mutant was investigated. After the optimization of the whole-cell conditions for the obtained mutant V18-N7, fed-batch conversion was carried out in a 5-L fermenter, and 44.13 g/L of PPA was synthesized with a conversion rate of 88%, which showed certain potential for industrial application. This study lays foundation for the industrial production of phenylpyruvic acid and also offers insights into the biosynthesis of other chemicals.
Escherichia coli/metabolism* ; Proteus mirabilis/genetics* ; Phenylpyruvic Acids/metabolism* ; Protein Engineering/methods* ; Recombinant Proteins/biosynthesis* ; Bacterial Proteins/metabolism*

L-tyrosine is one of the 20 amino acids that make up proteins and is an essential amino acid for mammals, often used as a nutritional supplement. The conventional methods for synthesizing L-tyrosine have some problems such as the production of many by-products, high requirements for production conditions, and environmental pollution. In this study, we designed and constructed a multi-enzyme cascade for the synthesis of L-tyrosine with alanine, glutamate, ammonium chloride, and phenol as substrates. Initially, the sources of glutamate oxidase, alanine aminotransferase, and tyrosine phenol lyase were screened and analyzed, which was followed by the identification of the rate-limiting enzyme in the reaction process. A colorimetric screening method was established, and the rate-limiting enzyme DbAlaA was engineered to enhance its activity by 40.0%. Subsequently, the reaction conditions, including temperature, pH, cell concentration, and surfactant and coenzyme dosages, were optimized. After optimization, the yield of L-tyrosine reached 9.93 g/L, with a alanine conversion rate of 54.90%. Finally, a feed-batch fermentation strategy was adopted, and the yield of L-tyrosine reached 56.07 g/L after 24 h, with a alanine conversion rate of 65.22%. This study provides a reference for the whole-cell catalytic synthesis of L-tyrosine and its industrialization.
Tyrosine/biosynthesis* ; Escherichia coli/metabolism* ; Tyrosine Phenol-Lyase/genetics* ; Multienzyme Complexes/metabolism* ; Fermentation

Objective To evaluate the detection performance of common strains and the antimicrobial binding capacity of four blood culture systems made by BMX-FA/N Plus,Zhengzhou Anto,Zhuhai DIER and Chongqing Zhongyuan.Methods According to the common pathogens of clinical bloodstream infections in Wuhan No.1 Hospital,ATCC standard isolates of Staphylococcus aureus,Streptococcus pneumoniae,Enterococcus faecalis,Escherichia coli,Pseudomonas aeruginosa,Haemophilus influenzae,Stenotrophomonas Maltophil,Bacteroides tenuis and Candida glabla were chosen to explore.Prefabricated bacterial suspension and sterile horse blood were injected into different brands of blood culture systems to simulate the blood samples of patients with non-antibiotic treatment and antibiotic treatment.Six commonly used clinical antibiotics,Imipenem,Piperacillin/Tazobactam,Cefoperazone/Sulbactam,Levofloxacin,Vancomycin and Micafungine,were added to the blood samples after simulated antibiotic treatment.The performance was evaluated by recording the positive bottles and the detection time of each brand culture systems within five days with and without antibiotic.Results In the absence of antibiotic,four blood culture systems showed 100％recovery on all of the pathogens.Staphylococcus aureus,Pseudomonas aeruginosa and Haemophilus influenzae were recovered earlier in DIER aerobic bottles than BMX-FA Plus aerobic bottles,and the differences were statistically significant(t=2.608,5.547,12.247,all P＜0.05).The time to detection of Staphylococcus aureus,Streptococcus pneumoniae,Enterococcus faecalis and Haemophilus influenzae in Zhongyuan anaerobic bottles was significantly faster than that of BMX-FA Plus anaerobic bottles,with an average shortening of 1.92～10.80 h,and the differences were statistically significant(t=30.187,5.367,33.068,24.855,all P＜0.05).When antibiotics added,BMX-FA Plus culture bottle showed 100％recovery to all the detecting pathogens with the peak concentration antibiotics except Cefoperazone/Sulbactam,while the recovery in Zhongyuan blood culture bottle also was 100％with peak concentration antibiotics of Piperacillin/Tazobactam,Levofloxacin and Micafunzin.The peak concentration of imipenem antibiotics in domestic bottles was only detected in Anto anaerobic bottles,with lower positive detection rate(66.7％)lower than that of BMX-FA Plus(100％)and a later detection,and the difference was statistically significant(t=-21.000,P=0.030).Conclusion In the absence of antibiotic interference,the positive detection rate of the above pathogens are the same for four blood culture systems,and the time to detection of DIER and Zhongyuan systems is shorter than that of BMX-FA/N Plus.In the presence of antibiotic interference,the detection ability to pathogens in BMX-FA/N Plus system is the best,followed by domestic Zhongyuan system.

At present, effective reconstruction of the integrity and functionality of damaged skin tissue remains an important medical problem in the field of wound repair. In recent years, the rapid development of nanozymes and tissue engineering scaffolds in the field of regenerative medicine has made it possible to develop new skin wound repair materials. Based on the process of skin wound repair and regeneration, this review briefly describes the nanozymes and its catalytic mechanism. At the same time, the common tissue engineering scaffolds loaded with nanozymes and their manufacturing strategies are introduced, the application of tissue engineering scaffolds loaded with nanozymes during the stages of anti-bacteria and anti-inflammation in the process of wound repair is summarized, and their future development direction is discussed.

At present, the implementation of Internet hospitals is still hampered by the double digital divide between doctors and patients. In order to develop Internet healthcare better, a Grade-A tertiary hospital has implemented digital governance for traditional healthcare system. The healthcare service chain has been reshaped by integrating online and offline health services and using multi-subject synergy and government-driven logic. Information barriers were broken down by strengthening hospital′s top-level design and implementing personalized follow-up visits for specialized diseases and live broadcast of health education. By establishing cross-regional specialist alliances and medical consortia oriented to patient needs, the time and space limitations of medical resources have been broken through. Digital governance practices have realized the two-way bridging of digital divide between doctors and patients, expanded the new mode of digital healthcare, and provided reference for the sustainable development of Internet hospitals.

Good anticoagulant biomaterials need good surface chemical properties, good mechanics performances and particularly good characteristics of biocompatibility, including tissue compatibility and hemocompatibility. In order to understand with greater clearness the anticoagulant biomaterial, we have to characterize them by different methods. In this paper, the approaches to assessing and displaying the characteristics of anticoagulant biomaterial are reviewed in three aspects, namely the surface chemical properties and structure, the mechanics performances the and the biocompatibility of anticoagulant biomaterial.
Anticoagulants ; Biocompatible Materials ; chemistry ; Blood Coagulation ; drug effects ; Humans ; Materials Testing ; Prostheses and Implants ; adverse effects ; Prosthesis Design ; Surface Properties