Immobilized enzyme-based enzyme electrode biosensors, characterized by high sensitivity and efficiency, strong specificity, and compact size, demonstrate broad application prospects in life science research, disease diagnosis and monitoring, etc. Immobilization of enzyme is a critical step in determining the performance (stability, sensitivity, and reproducibility) of the biosensors. Random immobilization (physical adsorption, covalent cross-linking, etc.) can easily bring about problems, such as decreased enzyme activity and relatively unstable immobilization. Whereas, directional immobilization utilizing amino acid residue mutation, affinity peptide fusion, or nucleotide-specific binding to restrict the orientation of the enzymes provides new possibilities to solve the problems caused by random immobilization. In this paper, the principles, advantages and disadvantages and the application progress of enzyme electrode biosensors of different directional immobilization strategies for enzyme molecular sensing elements by specific amino acids (lysine, histidine, cysteine, unnatural amino acid) with functional groups introduced based on site-specific mutation, affinity peptides (gold binding peptides, carbon binding peptides, carbohydrate binding domains) fused through genetic engineering, and specific binding between nucleotides and target enzymes (proteins) were reviewed, and the application fields, advantages and limitations of various immobilized enzyme interface characterization techniques were discussed, hoping to provide theoretical and technical guidance for the creation of high-performance enzyme sensing elements and the manufacture of enzyme electrode sensors.

T7 RNA polymerase (T7 RNAP) is one of the simplest known RNA polymerases. Its unique structural features make it a critical model for studying the mechanisms of RNA synthesis. This review systematically examines the static crystal structure of T7 RNAP, beginning with an in-depth examination of its characteristic “thumb”, “palm”, and “finger” domains, which form the classic “right-hand-like” architecture. By detailing these structural elements, this review establishes a foundation for understanding the overall organization of T7 RNAP. This review systematically maps the functional roles of secondary structural elements and their subdomains in transcriptional catalysis, progressively elucidating the fundamental relationships between structure and function. Further, the intrinsic flexibility of T7 RNAP and its applications in research are also discussed. Additionally, the review presents the structural diagrams of the enzyme at different stages of the transcription process, and through these diagrams, it provides a detailed description of the complete transcription process of T7 RNAP. By integrating structural dynamics and kinetics analyses, the review constructs a comprehensive framework that bridges static structure to dynamic processes. Despite its advantages, T7 RNAP has a notable limitation: it generates double-stranded RNA (dsRNA) as a byproduct. The presence of dsRNA not only compromises the purity of mRNA products but also elicits nonspecific immune responses, which pose significant challenges for biotechnological and therapeutic applications. The review provides a detailed exploration of the mechanisms underlying dsRNA formation during T7 RNAP catalysis, reviews current strategies to mitigate this issue, and highlights recent progress in the field. A key focus is the semi-rational design of T7 RNAP mutants engineered to minimize dsRNA generation and enhance catalytic performance. Beyond its role in transcription, T7 RNAP exhibits rapid development and extensive application in fields, including gene editing, biosensing, and mRNA vaccines. This review systematically examines the structure-function relationships of T7 RNAP, elucidates the mechanisms of dsRNA formation, and discusses engineering strategies to optimize its performance. It further explores the engineering optimization and functional expansion of T7 RNAP. Furthermore, this review also addresses the pressing issues that currently need resolution, discusses the major challenges in the practical application of T7 RNAP, and provides an outlook on potential future research directions. In summary, this review provides a comprehensive analysis of T7 RNAP, ranging from its structural architecture to cutting-edge applications. We systematically examine: (1) the characteristic right-hand domains (thumb, palm, fingers) that define its minimalistic structure; (2) the structure-function relationships underlying transcriptional catalysis; and (3) the dynamic transitions during the complete transcription cycle. While highlighting T7 RNAP’s versatility in gene editing, biosensing, and mRNA vaccine production, we critically address its major limitation—dsRNA byproduct formation—and evaluate engineering solutions including semi-rationally designed mutants. By synthesizing current knowledge and identifying key challenges, this work aims to provide novel insights for the development and application of T7 RNAP and to foster further thought and progress in related fields.

Aim To investigate the effect of homocys-teine(Hcy)on the permeability of pulmonary micro-vascular endothelial cells(PMVECs)and the role and mechanism of RASAL1.Methods CBS+/-mice were fed a high methionine diet(HMD)for 16 weeks to replicate an animal model of hyperhomocysteinemia(HHcy).HE staining was used to observe the changes in lung tissue structure.qRT-PCR was used to detect the levels of RASAL1 and DNMT1 mRNA in lung tis-sue.Western blot was used to detect the expression of RASAL1,DNMT1,ZO-1,and VE cadherin proteins.Methylation specific PCR was used to detect methyla-tion in the RASAL1 promoter region.PMVECs were transfected with Ad-RASAL1 to detect the expression of ZO-1 and VE cadherin.The si-DNMT1 interference fragment was transfected into PMVECs,and the ex-pression of the RASAL1 was detected by qRT-PCR and Western blot.Results Serum Hcy level of HMD mice was significantly raised,and HE staining showed severe structural disorder in lung tissue.The expres-sion of RASAL1,ZO-1,and VE cadherin was de-creased,while the expression of DNMT1 was in-creased.The degree of methylation in the RASAL1 promoter region was raised.The expression of ZO-1 and VE cadherin increased after PMVECs were trans-fected with Ad-RASAL1.After knocking down DN-MT1,RASAL1 expression was increased.Conclusion Hcy can increase the permeability of PMVECs,and its mechanism is related to the upregulation of RASAL1 methylation level.

Aim To explore the antimicrobial activity of a novel R-type phage tail-like bacteriocin(PTLB)secreted by Enterobacter cloacae SHAMU191747 a-gainst methicillin-resistant Staphylococcus aureus(MR-SA).Methods Antagonistic activity of E.cloacae SHAMU191747 against MRS A was detected by LB agar plate antagonistic test and LB broth micro-fermentation test.The crude extract of fermentation supernatant of E.cloacae SHAMU191747 was prepared by ultra-high-speed centrifugation and density gradient centrifuga-tion.The novel R-type PTLB in the crude extract was detected by transmission electron microscopy.The an-timicrobial activity of the crude extract against MRSA was verified by LB agar plate spot-seeding method.The molecular weight of the novel R-type PTLB was detected by SDS-PAGE electrophoresis.Results E.cloacae SHAMU191747 secreted a novel R-type PTLB,and had a strong antagonistic effect on MRSA.The no-vel R-type PTLB had a molecular weight of approxi-mately 35 ku and could efficiently kill MRSA.The physical dimensions of its tail-sheath-uncontracted functional molecules were(142.7±4.3)×(13.8±0.6)nm,and those of the tail-sheath-contracted non-functional molecules were(57.7±1.2)×(20.8±1.5)nm.Conclusions The novel R-type PTLB pro-duced by E.cloacae SHAMU191747 can efficiently kill MRSA,and has the potential to be developed into a novel antimicrobial drug with great prospects for clini-cal application.

This study aims to obtain two whole-genome sequences of enzootic nasal tumor virus of goats(ENTV-2)from Anhui Province and analyzed the genetic diversity of ENTV-2 gene.Nasal secretion samples and blood samples of six goats with enzootic nasal adenocarcinoma(ENA)were collected from a goat farm in Anhui Province.The total RNA was extracted by the TRIzol method.The DNA interference was removed by the two-step reverse transcription.The ENTV-2 was detec-ted by PCR.Then,two positive samples were selected and five pairs of primers were used to ampli-fy the whole-genome sequences of ENTV-2.After sequencing and splicing,two sequences were up-loaded to the database for comparative analysis with the sequences in the NCBI database.Finally,the genetic evolution tree was constructed.ENTV-2 was detected in the nasal secretion samples,but not in the blood of the six ENA goats.The ENTV-2 genes were approximately 7 400 bp in length,named ENTV-2AH1(DDBJ accession no.:LC762616)and ENTV-2AH2(DDBJ accession no.:LC762617),respectively.Two sequences showed 99.2％and 99.1％homology with the Fujian strain(ENTV-2FJ)and Guangxi strain(ENTV-2-DA0),respectively.They were in the same evo-lutionary branch.In this study,two whole-genome sequences of ENTV-2 were obtained in Anhui for the first time,which can help to further study the genetic diversity of ENTV-2 in China.

Objective:To analyze the immune reconstitution after BTKi treatment in patients with chronic lymphocytic leukemia(CLL).Methods:The clinical and laboratorial data of 59 CLL patients admitted from January 2017 to March 2022 in Fujian Medical University Union Hospital were collected and analyzed retrospectively.Results:The median age of 59 CLL patients was 60.5(36-78).After one year of BTKi treatment,the CLL clones(CD5+/CD19+)of 51 cases(86.4％)were significantly reduced,in which the number of cloned-B cells decreased significantly from(46±6.1)× 109/L to(2.3±0.4)× 109/L(P=0.0013).But there was no significant change in the number of non-cloned B cells(CD19+minus CD5+/CD19+).After BTKi treatment,IgA increased significantly from(0.75±0.09)g/L to(1.31±0.1)g/L(P＜0.001),while IgG and IgM decreased from(8.1±0.2)g/L and(0.52±0.6)g/L to(7.1±0.1)g/L and(0.47±0.1)g/L,respectively(P＜0.001,P=0.002).BTKi treatment resulted in a significant change in T cell subpopulation of CLL patients,which manifested as both a decrease in total number of T cells from(2.1±0.1)× 109/L to(1.6±0.4)× 109/L and NK/T cells from(0.11±0.1)× 109/L to(0.07±0.01)× 109/L(P=0.042,P=0.038),both an increase in number of CD4+cells from(0.15±6.1)× 109/L to(0.19±0.4)× 109/L and CD8+cells from(0.27±0.01)× 109/L to(0.41±0.08)× 109/L(both P＜0.001).BTKi treatment also up-regulated the expression of interleukin(IL)-2 while down-regulated IL-4 and interferon(IFN)-γ.However,the expression of IL-6,IL-10,and tumor necrosis factor(TNF)-α did not change significantly.BTKi treatment could also restored the diversity of TCR and BCR in CLL patients,especially obviously in those patients with complete remission(CR)than those with partial remission(PR).Before and after BTKi treatment,Shannon index of TCR in patients with CR was 0.02±0.008 and 0.14±0.001(P＜0.001),while in patients with PR was 0.01±0.03 and 0.05±0.02(P＞0.05),respectively.Shannon index of BCR in patients with CR was 0.19±0.003 and 0.33±0.15(P＜0.001),while in patients with PR was 0.15±0.009 and 0.23±0.18(P＜0.05),respectively.Conclusions:BTKi treatment can shrink the clone size in CLL patients,promote the expression of IgA,increase the number of functional T cells,and regulate the secretion of cytokines such as IL-2,IL-4,and IFN-γ.BTKi also promote the recovery of diversity of TCR and BCR.BTKi treatment contributes to the reconstitution of immune function in CLL patients.

Objective:To explore the quality of working life and its related factors in patients with breast canc-er returned to work after surgery.Methods:A total of 316 patients with breast cancer who had returned to work af-ter surgery were selected,and they were investigated with the General information questionnaire,Quality of Working Life Questionnaire for Cancer Survivors(QWLQ-CS),Breast Cancer Survivors Resilience Scale(BCRS)and Strat-egies Used by People to Promote Health(SUPPH).Multivariate linear regression analysis was used to analyze the related factors of quality of working life.Results:Multiple linear regression analysis showed that the QWLQ-CS score was positively correlated with education level,per capita monthly income of family,work pattern,return to work time,personal protection and social protection score of BCRS,and positive attitude scores of SUPPH(β=1.05,1.23,2.26,0.69,0.95,1.00,0.13),while negatively correlated with the complications(β=-3.83).Con-clusion:The quality of working life for patients with breast cancer after surgery needs to be improved after they re-turn to work,psychological resilience and self-efficacy are positively correlated with quality of working life.

Objective To develop a medical positive pressure protective suit for long-voyage aeromedical evacuation to realize protective isolation and drinking water and energy supply for medical personnel during long-voyage aeromedical evacuation of respiratory infectious disease patients.Methods The medical positive pressure protective suit had a one-piece structure,with its main part made of hydroentangled non-woven fabric,head and forebreast parts made of amorphous polyethylene terephthalate anti-fog material and wearing-and taking-off parts sealed with zipper and autohesion,which was equipped with a portable positive pressure air supply device,an airborne centralized positive pressure air supply device and a monitoring and warning device.The portable positive pressure air supply device was fixed in the back of the suit at the waist,the airborne device was made by modifying the commercially available positive pressure air supply fan,and the monitoring and warning device monitored the air supply volume of the fan,the battery power and the pressure inside the suit.Results The suit behaved well in protection,clean fresh air supply without time limitation and facilitating hydration and energy replenishment of medical personnel by forming three activity spaces.Conclusion The suit developed can continuously provide a clean and comfortable microenvironment,meeting the requirements of medical personnel for protection and hydration and energy replenishment during long-voyage aeromedical evacuation.[Chinese Medical Equipment Journal,2024,45(11):113-116]

Objective To investigate whether the mechanism of action of electro-acupuncture in MPTP-induced Parkinson's disease(PD)model mice is related to neuroinflammation mediated by the forkhead box protein O1(FoXO1)/NOD-like receptor heat protein structural domain-related protein 3(NLRP3)signaling pathway.Methods Thirty mice were randomly divided into control(Ctrl),model(MPTP),and electro-acupuncture(EA)groups,with 10 mice in each group.PD mouse model was prepared by intraperitoneal injection of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP)30 mg/(kg·d)for 7 d consecutively.Behavioral changes of mice were observed by open field and pole-climbing experiments;α-synuclein(α-syn)expression in the substantia nigra of the midbrain was observed by immunohistochemistry;FoXO1,NLRP3,ASC,and Caspase-1 were deter-mined by Western blot;and the expression of inflammatory factors such as tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β)was detected by ELISA in the substantia nigra of the midbrain.Results Mice in the MPTP group showed significant behavioral deficits,with a significant increase in midbrain substantia nigra α-syn(P＜0.01),elevated levels of Iba-1,FoXO1,NLRP3,ASC,and Caspase-1 proteins(P＜0.05 and P＜0.01,respectively),and a significant increase in the levels of TNF-α,and IL-1β(P＜0.01).The treatment of EA was effective in improve the dyskinesia of PD mice,improve motor balance and coordination(P＜0.01),reduce the abnormal aggregation of α-syn(P＜0.01),reduce the abnormal activation of microglia,and alleviate neuroinflammation(P＜0.05).Conclusion Electro-acupuncture may play a protective role in PD by modulating FoXO1/NLRP3 pathway-mediated neuroinflammation.

Objective To analyze the epidemiological and clinical characteristics of patients with monkeypox.Methods Data of 17 patients with monkeypox hospitalized in a hospital in Nanning City from July to October 2023 were collected retrospectively.The epidemiological history,clinical manifestations,laboratory examinations,treat-ment and prognosis were analyzed and summarized.Results All 17 patients were male,with a median age of 28 years old.Fifteen(88.2％)patients were men who had sex with men(MSM)within 21 days prior to onset.Major clinical manifestations were rash and fever.Rashes distributed mainly in the anus,perineum and genitals(82.4％),followed by the trunk and limbs(52.9％),head and face(35.3％),while soles and palms were rare.Some patients had swollen inguinal lymph nodes.All patients were discharged from hospital after improvement,with an average hospital stay of 7 days.Conclusion The monkeypox epidemic in Nanning area of Guangxi occurs mainly in MSM population,with fever and rashes as the major symptoms.All patients have mild disease and good prognosis.