The rapid development of the biomanufacturing industry has put an increasing demand for interdisciplinary talents equipped with strong bioengineering design capabilities. However, the conventional teaching model of the AutoCAD Design course fails to effectively support the cultivation of high-quality bioengineering professionals, as students commonly face challenges such as weak engineering application ability, limited innovation and practical skills, and poor alignment with industry needs. To address these issues and enhance students' integrated design and engineering innovation capabilities, we developed a systematic teaching reform framework known as the "one core and five dimensions" model according to the student-centered and competence-oriented educational philosophy. This framework drives comprehensive reforms in teaching contents and teaching methods through five dimensions: curriculum system optimization, online and offline teaching integration, production-education collaboration, incorporation of ideological and political elements, and the integration of competitions with teaching. Since the reform measures were implemented, students have exhibited significantly improvements in operational skills, practical ability, and innovation awareness, which indicate that the overall teaching effectiveness of this course has continued to strengthen. This study offers an effective path for reforming the AutoCAD Design course in bioengineering programs and provides valuable insights for optimizing the talent competency structure in the biomanufacturing field.

Gibberellic acid (GA3) is a crucial phytohormone involved in plant growth regulation with widespread applications in agriculture and other fields. In this study, GA3 synthesis in an industrial GA3 producer F. fujikuroi strain FF00 was improved with a strategy by reprogramming the regulatory network and overexpressing key genes. Three positive regulators AreA (the nitrogen regulator), Lae1 (the velvet complex component), and Hat1 (the histone modification regulator) were identified by screening regulators affecting GA3 biosynthesis. Mutant FF18-4 with GA3 titer of 2.61g/L in shake-flask fermentation was constructed by co-overexpressing AreA, Lae1, and Hat1 in strain FF00. Comparative transcriptomic analysis revealed that reprogramming of the regulatory network resulted in the down-regulation of two key genes (Ggs2 and Cps/Ks) in the GA3 biosynthetic pathway which reduced the supply of GA3 precursor geranylgeranyl pyrophosphate. Ggs2 and Cps/Ks genes were further co-overexpressed to enhance metabolic flux for GA3 biosynthesis. The GA3 titer of the resulting mutant FF19-5 reached 2.73g/L, which was 49.2% higher than that of strain FF00. Therefore, the GA3 biosynthesis of strain FF00 was significantly improved by metabolic network reprogramming and metabolic balance. The established strategy provided the basis for GA3 over-producer construction and may be helpful for the synthesis of other chemicals with microbial cell factories.

A robust and reusable Rhizomucor miehei lipase (RML) biocatalyst was prepared by integrating bio-imprinting with resin immobilization. RML was co-imprinted with lauric acid and polyethylene glycol (PEG600) followed by immobilization on an amine-functionalized epoxy resin (ES-103B), resulting in AE@RML-LA+PEG600. After immobilization process optimization, the AE@RML-LA + PEG 600 was verified by CD, SEM, FTIR, and XRD analysis. AE@RML-LA+PEG600 exhibited improved thermal, pH, and organic solvent stability compared with free RML. Under optimized conditions (10 h, 62 °C, palmitic-acid-enriched palm stearin/oleic acid = 1:10 mol/mol, 10 wt

Gibberellic acid (GA3) is an important plant growth regulator with rising agricultural demand, yet its industrial production via submerged fermentation using Fusarium fujikuroi remains limited by low space-time yield. In this study, a high-yield mutant was generated through atmospheric and room-temperature plasma (ARTP) and ethyl methanesulfonate (EMS) mutagenesis, coupled with lovastatin-based screening. Process optimization, including early nitrogen supplementation, late-stage pH control, and a two-stage temperature-agitation strategy, further enhanced GA3 synthesis. The mutant (strain 9) achieved a GA3 titer of 2.483 g/L in a 5-L bioreactor, a 33.38

The biocatalytic production of D-tagatose, a valuable low-calorie sweetener, is often hindered by poor enzyme stability and limited reusability. To address this issue, a thermostable tagatose-4-epimerase (TpT4Ease) from Thermotoga petrophila was crosslinked with glutaraldehyde followed by immobilization on an amino-functionalized macroporous resin (LX-1000HA), resulting in TpT4Ease@LX-1000HA-GA.The effective synthesis of TpT4Ease@LX-1000HA-GA was verified by CD and SEM after optimization of the immobilization procedure. Kinetic analysis revealed that the conformational constraint within the porous support slightly compromised intrinsic turnover but effectively prevented thermal unfolding. Consequently, TpT4Ease@LX-1000HA-GA displayed improved thermal and storage stability, with its half-life at 70 °C extended to 15.4 h compared to 3.3 h for the free enzyme. Under optimized continuous-flow conditions (200 g/L fructose, 70 °C, 0.5 mL/min), TpT4Ease@LX-1000HA-GA achieved an efficient bioconversion of D-fructose to D-tagatose with a yield of 17.3

This study aimed to characterize the G protein-coupled receptor FfGpr1-Gα-AC transduction pathway and its role in regulating gibberellin (GA) metabolism in Fusarium fujikuroi. By constructing Ffgpr1 deletion and constitutively activated FfG2Q204 L mutants, we found that glucose-induced cAMP (cyclic Adenosine MonoPhosphate) synthesis was abolished in Ffgpr1Δ. Bimolecular fluorescence complementation confirmed membrane-localized FfGpr1-FfG2 and FfG2-AC interactions, with intensified fluorescence at septa. Fermentation assays revealed opposing GA3 yields: Ffgpr1Δ produced 21% less GA3 than the wild type, whereas FfG2Q204 L increased yield by 17%. qPCR（quantitative real-time PCR） analysis demonstrated that Ffgpr1Δ upregulated FfCPS/KS, FfP450-2, and FfP450-3 transcription by 6-8-fold while downregulating FfDES by 82%, whereas FfG2Q204 L induced a 6-fold increase in FfCPS/KS mRNA level. Strikingly, FfDES overexpression in Ffgpr1Δ restored GA3 production to wild-type levels but led to GA7 accumulation and suppressed FfP450-3 upregulation, suggesting feedback-regulated metabolic constraints. Glucose-induced cAMP production required FfGpr1. FfGpr1-FfG2 and FfG2-AC interacted on the cell membrane, with enhanced co-localization at the septal region. The FfGpr1-Gα-AC pathway significantly affected GA yield, with complex and noteworthy regulation of GA cluster gene expression.

S-adenosyl-L-methionine (SAM) is the only injectable drug among the hepatoprotective and choleretic drugs, which has remarkable efficacy and is favored by hepatopaths. The demand for SAM is constantly increasing in clinical settings. Therefore, many efforts have been made to increase SAM biosynthesis from L-methionine and ATP in Saccharomyces cerevisiae. This study aimed to construct a stable and high-accumulating SAM industrial strain through successive ultraviolet irradiation (UV) mutations coupled with three resistant (ethionine, nystatin, and cordycepin, respectively) screening procedures and metabolic engineering strategies. Following multiple UV mutagenesis, a higher production mutant strain ZJT15-33 was successfully obtained. In addition, the recombinant strain spe2 triangle-PPX1 was derived from ZJT15-33 by deleting the SPE2 and overexpressing the PPX1, resulting in a 2.5-fold enhanced ATP accumulation, which promoted the synthesis of 2.41 g/L SAM in the shakeflask, representing an 11.4-fold enhancement over the original strain (0.21 g/L). Furthermore, 11.65 g/L SAM was accumulated with 113 mg/g DCW SAM content in a 5-L fermenter at 96 h, marking a 36.57 % increase compared to strain ZJT15-33 (8.53 g/L). These results indicated that UV mutagenesis combined with PPX1 overexpression could effectively improve SAM synthesis in S. cerevisiae, providing a feasible approach for developing highly SAM industrial production.

S-Adenosyl-L-methionine (SAM), an important biochemical substance, is experiencing increased demand due to its versatile applications in the medical field. Although Saccharomyces cerevisiae has been adapted as a promising platform for SAM production, the issue of SAM productivity is still very interesting. In this study, a systematic approach that consists of genome shuffling, genetic engineering, and fermentation process optimization strategies was established. Firstly, haploids of mutant T11-1 and P15-33 were prepared and treated with ultraviolet irradiation (UV) and atmospheric and room temperature plasma (ARTP) compound mutagenesis, which aids to increase SAM production to 1.25, 1.34 g/L, respectively. Genome shuffling was performed with cycloheximide-resistant protoplast of mutant T5-8-1 and hygromycin B-resistant protoplasts of mutant P4-2-4. The resulting fusant TRP60, with a SAM titer of 1.36 g/L, was subjected to transcriptome analysis. Subsequently, hemoglobin from vitreoscilla has been introduced into fusant TRP60 to increase ATP supply and then strengthening the synthesis pathway of SAM, which significantly elevated SAM titer to 2.06 g/L. Finally, the SAM titer of fusant TRP60-vgb-SAM2 reached 14.22 g/L in a 5 L bioreactor with an optimized fed-batch fermentation. Therefore, fusant TRP60-vgb-SAM2 may serve as a candidate for industrial production of SAM.

S-adenosyl-L-methionine (SAM) is an important compound with significant pharmaceutical and nutraceutical applications. Currently, microbial fermentation is dominant in SAM production, which remains challenging due to its complex biosynthetic pathway and insufficient precursor availability. In this study, a multimodule engineering strategy based on CRISPR/Cas9 was established to improve the SAM productivity of Saccharomyces cerevisiae . This strategy consists of (1) improving the growth of S. cerevisiae by overexpressing the hxk2 gene; (2) enhancing the metabolic flux toward SAM synthesis by upregulating the expression of the aat1, met17 , and sam2 genes and weakening the synthesis pathway of L-threonine; (3) elevating precursor ATP synthesis by introducing the vgb gene; (4) blocking the SAM degradation pathway by knocking out the sah1 and spe2 genes. The SAM titer of the resulting mutant AU18 reached 1.87 g/L, representing an increase of 227.67% compared to the parental strain. With optimal medium, SAM titer of mutant AU18 reached 2.46 g/L in flask shake fermentation. The SAM titer of mutant AU18 further reached 13.96 g/L after 96 h incubation with a continuous L-Met feeding strategy in a 5 L fermenter. Therefore, with comprehensive optimization of both synthesis and degradation pathways of SAM, a multimodule strategy was established, which significantly elevated the SAM production of S. cerevisiae . This laid a foundation for the construction of hyperproducer for SAM and other valuable amino acids or chemicals.

AIMS:This study aimed to characterize the G protein-coupled receptor FfGpr1-Gα-AC transduction pathway and its role in regulating gibberellin (GA) metabolism in Fusarium fujikuroi. METHODS AND RESULTS:By constructing Ffgpr1 deletion and constitutively activated FfG2Q204 L mutants, we found that glucose-induced cAMP (cyclic Adenosine MonoPhosphate) synthesis was abolished in Ffgpr1Δ. Bimolecular fluorescence complementation confirmed membrane-localized FfGpr1-FfG2 and FfG2-AC interactions, with intensified fluorescence at septa. Fermentation assays revealed opposing GA3 yields: Ffgpr1Δ produced 21% less GA3 than the wild type, whereas FfG2Q204 L increased yield by 17%. qPCR（quantitative real-time PCR） analysis demonstrated that Ffgpr1Δ upregulated FfCPS/KS, FfP450-2, and FfP450-3 transcription by 6-8-fold while downregulating FfDES by 82%, whereas FfG2Q204 L induced a 6-fold increase in FfCPS/KS mRNA level. Strikingly, FfDES overexpression in Ffgpr1Δ restored GA3 production to wild-type levels but led to GA7 accumulation and suppressed FfP450-3 upregulation, suggesting feedback-regulated metabolic constraints. CONCLUSION:Glucose-induced cAMP production required FfGpr1. FfGpr1-FfG2 and FfG2-AC interacted on the cell membrane, with enhanced co-localization at the septal region. The FfGpr1-Gα-AC pathway significantly affected GA yield, with complex and noteworthy regulation of GA cluster gene expression.

S-adenosyl-L-methionine (SAM) plays pivotal roles in various physiological processes. With its increasing application in the treatment of diseases such as liver disease, depression, osteoarthritis and Alzheimer's, interest in SAM production aroused. Currently, Saccharomyces cerevisiae is the main industrial producer of SAM. With the surge in demand for SAM, improving the SAM biosynthesis is of importance. In this study, a multimodule engineering strategy was employed to improving SAM production: 1) Enhancing the gene expression of the sulfur assimilation pathway; 2) Strengthening the metabolic flux of the SAM synthesis pathway; 3) Weakening the SAM degradation pathway; 4) Increasing ATP supply. The resulting engineered mutant SC06 (S. cerevisiae CEN.PK2-1C Delta gal80::T-cyc1-sam2-P-gal1-P-gal10-met14-T-adh1, Delta lsc2::T-cyc1-hom6-P-gal1-P-gal10-met6-T-adh1, Delta sah1 Delta mls1) displayed the highest SAM titer of 240.86 mg/L, which was 10.22-fold increase compared with the original strain. With optimized conditions, the SAM titer of mutant SC06 in shake flask fermentation reached 473.02 mg/L with a specific yield of 127.18 mg/g dry cell weight (DCW). In a 5 L fermenter with fed-batch fermentation, the maximal SAM yield of mutant SC06 reached 1.25 g/L with a specific yield of 166.67 mg/g DCW after 58 h cultivation. Therefore, the established metabolic engineering strategies displayed promising efficiency in improving the SAM productivity of S. cerevisiae CEN.PK2-1C, which may provide a useful tool for the improvement of SAM-producing strains.

With potent herbicidal activity, biocatalysis synthesis of l-glufosinate has drawn attention. In present research, NAP-Das2.3, a deacetylase capable of stereoselectively resolving N-acetyl-l-glufosinate to l-glufosinate mined from Arenimonas malthae, was heterologously expressed and characterized. In Escherichia coli, NAP-Das2.3 activity only reached 0.25 U/L due to the formation of inclusive bodies. Efficient soluble expression of NAP-Das2.3 was achieved in Pichia pastoris. In shake flask and 5 L bioreactor fermentation, NAP-Das2.3 activity by recombinant P. pastoris reached 107.39 U/L and 1287.52 U/L, respectively. The optimum temperature and pH for N-acetyl-glufosinate hydrolysis by NAP-Das2.3 were 45 °C and pH 8.0, respectively. The Km and Vmax of NAP-Das2.3 towards N-acetyl-glufosinate were 25.32 mM and 19.23 μmol mg−1 min−1, respectively. Within 90 min, 92.71

S-adenosyl-l-methionine (SAM) is ubiquitous in living organisms and plays important roles in transmethylation, transsulfuration and transamination in organisms. Due to its important physiological functions, production of SAM has attracted increasing attentions. Currently, researches on SAM production mainly focus on microbial fermentation, which is more cost-effective than that of the chemical synthesis and the enzyme catalysis, thus easier to achieve commercial production. With the rapid growth in SAM demand, interests in improving SAM production by developing SAM hyper-producing microorganisms aroused. The main strategies for improving SAM productivity of microorganisms include conventional breeding and metabolic engineering. This review summarizes the recent research progress in improving microbial SAM productivity to facilitate further improving SAM productivity. The bottlenecks in SAM biosynthesis and the solutions were also addressed.

新经济形势给高等学校生物工程专业的"新工科"建设提出了新的要求.在生物工程专业课程教学过程中,与大型发酵设备及发酵工厂相关的教学手段受到诸多限制.针对此类问题,开展计算机虚拟仿真实验项目的开发与建设具有重要意义.简述了国内大型发酵车间虚拟仿真实验建设的状况,介绍了国家虚拟仿真实验教学课程共享平台上的 2 个典型的 3D 大型发酵工厂虚拟仿真实验.结合建设中的阿卡波糖发酵车间 3D 虚拟仿真实验的四大模块内容及教学实践成果的介绍,探讨了大型发酵工厂虚拟仿真实验课程建设尚存在的问题及今后的发展方向.

Fusarium fujikuroi is the microorganism that used for industrial production of gibberellic acids (GAs), a commercially very important plant hormone. A stable, high-yielding F. fujikuroi strain is essential for efficient bio-production. Although, there is a quick development in molecular tools and metabolic engineering research for this microbe in the past decade, the current industrially applying strains are mostly obtained from numerous rounds of mutagenesis and screening, while the industry is still dedicating in strain improvement work based mostly on this method. However, after over half a century's effort, the yield is still maintained at a low level today. We designed an efficient strategy for strain improvement by amphotericin B resistance prescreening and evolutionary engineering. A superior strain was obtained with over 25% increased yield after testing merely 640 isolates. The superior strain was later confirmed to be genetically very stable for GA production. Minimum inhibitory concentration (MIC) assay verified its increased resistance to amphotericin B, whose target ergosterol shares a same precursor with GAs. Decreased ergosterol accumulation indicated the increased GA3 synthesis accompanied with attenuation of metabolic branch flux. In the end, we found the increased GA3 titer of the superior strain also accompanied with faster growing biomass.

S-adenosyl-l-methionine (SAM), a vital physiologically active substance in living organisms, is produced by fermentation over Saccharomyces cerevisiae . The main limitation in SAM production was the low biosynthesis ability of SAM in S. cerevisiae . The aim of this work is to breed an SAM-overproducing mutant through UV mutagenesis coupled with high-throughput selection. Firstly, a high-throughput screening method by rapid identification of positive colonies was conducted. White colonies on YND medium were selected as positive strains. Then, nystatin/sinefungin was chosen as a resistant agent in directed mutagenesis. After several cycles of mutagenesis, a stable mutant 616–19-5 was successfully obtained and exhibited higher SAM production (0.41 g/L vs 1.39 g/L). Furthermore, the transcript levels of the genes SAM2 , ADO1 , and CHO2 involved in SAM biosynthesis increased, while ergosterol biosynthesis genes in mutant 616–19-5 significantly decreased. Finally, building on the above work, S. cerevisiae 616–19-5 could produce 10.92 ± 0.2 g/L SAM in a 5-L fermenter after 96 h of fermentation, showing a 2.02-fold increase in the product yield compared with the parent strain. Paving the way of breeding SAM-overproducing strain has improved the good basis for SAM industrial production.

The formation of calcium carbonate crystals in concrete by urease-producing bacteria is not understood fully. In this study, a Lysinibacillus boronitolerans strain with a high urease activity was isolated and used to analyze the counts and sizes of the crystals and the relationship with time.

Acarbose is widely used as α-glucosidase inhibitor in the treatment of type Ⅱ diabetes. Actinoplanes sp. is used for industrial production of acarbose. As a secondary metabolite, the biosynthesis of acarbose is quite complex. In addition to acarbose, a few acarbose structural analogs are also accumulated in the culture broth of Actinoplanes sp., which are hard to remove. Due to lack of systemic understanding of the biosynthesis and regulation mechanisms of acarbose and its structural analogs, it is difficult to eliminate or reduce the biosynthesis of the structural analogs. Recently, the advances in omics technologies and molecular biology have facilitated the investigations of biosynthesis and regulatory mechanisms of acarbose and its structural analogs in Actinoplanes sp.. The genes involved in the biosynthesis of acarbose and its structural analogs and their regulatory mechanism have been extensively explored by using bioinformatics analysis, genetic manipulation and enzymatic characterization, which is summarized in this review.

To improve S-Adenosyl-L-methionine (a compound with important physiological functions, SAM) production, atmospheric and room temperature plasma and ultraviolet-LiCl mutagenesis were carried out with Saccharomyces cerevisiae strain ZY 1-5. The mutants were screened with ethionine, L-methionine, nystatin and cordycepin as screening agents. Adaptive evolution of a positive mutant UV6-69 was further performed by droplet microfluidics cultivation with ethionine as screening pressure. After adaptation, mutant T11-1 was obtained. Its SAM titer in shake flask fermentation reached 1.31 g/L, which was 191% higher than that of strain ZY 1-5. Under optimal conditions, the SAM titer and biomass of mutant T11-1 in 5 L bioreactor reached 10.72 g/L and 105.9 g dcw/L (142.86% and 34.22% higher than those of strain ZY 1-5), respectively. Comparative transcriptome analysis between strain ZY 1-5 and mutant T11-1 revealed the enhancements in TCA cycle and gluconeogenesis/glycolysis pathways as well as the inhibitions in serine and ergosterol synthesis of mutant T11-1. The elevated SAM synthesis of mutant T11-1 may attribute to the above changes. Taken together, this study is helpful for industrial production of SAM.

As a strategic emerging industry of China, the biotechnology industry develops rapidly in recent years, which significantly increased the demand for creative and capable talents. As a core curriculum of bioengineering specialty, biotechnology equipment plays an important role in fostering such talents. To address the problems in biotechnology equipment course teaching such as limited equipment availability, limited engineering practice, and lack of learning motivations, curriculum reform and optimization were performed based on curriculum resource development, virtual reality-physical combined engineering training, and boosting learning motivations. The optimized teaching contents focus on fostering morality, intelligence, and creative practice abilities by connecting new requirements of social development, introducing new progress in biotechnology research, as well as new practices in research and development (R & D). Measures such as teaching methods innovation, assessment and evaluation methods optimization, cutting-edge R & D progress, diverse resources integration, and online-offline combined teaching, were developed to boost the learning motivation and foster the innovation competence of students. By above exploration and practice, the practice and innovation competence of students were significantly enhanced.