Major royal-jelly proteins intake modulates immune functions and gut microbiota in mice

In this study,we investigated the effects of major royal jelly proteins(MRJPs)on the estrogen,gut microbiota,and immunological responses in mice.Mice given 250 or 500 mg/kg,not 125 mg/kg of MRJPs,enhanced the proliferation of splenocytes in response to mitogens.The splenocytes and mesenteric lymphocytes activated by T-cell mitogens(Con A and anti-CD3/CD28 antibodies)released high levels of IL-2 but low levels of IFN-γand IL-17A.The release of IL-4 was unaffected by MRJPs.Additionally,splenocytes and mesenteric lymphocytes activated by LPS were prevented by MRJPs at the same dose as that required for producing IL-1βand IL-6,two pro-inflammatory cytokines.The production of IL-1β,IL-6,and IFN-γwas negatively associated with estrogen levels,which were higher in the MRJP-treated animals than in the control group.Analysis of the gut microbiota revealed that feeding mice 250 mg/kg of MRJPs maintained the stability of the natural intestinal microflora of mice.Additionally,the LEf Se analysis identified biomarkers in the MRJP-treated mice,including Prevotella,Bacillales,Enterobacteriales,Gammaproteobacteria,Candidatus_Arthromitus,and Shigella.Our results showed that MRJPs are important components of royal jelly that modulate host immunity and hormone levels and help maintain gut microbiota stability.

Essential proteins identification method based on four-order distances and subcellular localization information

Essential proteins are inseparable in cell growth and survival. The study of essential proteins is important for understanding cellular functions and biological mechanisms. Therefore, various computable methods have been proposed to identify essential proteins. Unfortunately, most methods based on network topology only consider the interactions between a protein and its neighboring proteins, and not the interactions with its higher-order distance proteins. In this paper, we propose the DSEP algorithm in which we integrated network topology properties and subcellular localization information in protein–protein interaction(PPI) networks based on four-order distances, and then used random walks to identify the essential proteins. We also propose a method to calculate the finite-order distance of the network, which can greatly reduce the time complexity of our algorithm. We conducted a comprehensive comparison of the DSEP algorithm with 11 existing classical algorithms to identify essential proteins with multiple evaluation methods. The results show that DSEP is superior to these 11 methods.

Prognosis value of heat-shock proteins in esophageal and esophagogastric cancer:A systematic review and meta-analysis

BACKGROUND Heat shock proteins(HSPs)are molecular chaperones that play an important role in cellular protection against stress events and have been reported to be overex-pressed in many cancers.The prognostic significance of HSPs and their regulatory factors,such as heat shock factor 1(HSF1)and CHIP,are poorly understood.AIM To investigate the relationship between HSP expression and prognosis in esophageal and esophagogastric cancer.METHODS A systematic review was conducted in accordance with PRISMA recommend-ations(PROSPERO:CRD42022370653),on Embase,PubMed,Cochrane,and LILACS.Cohort,case-control,and cross-sectional studies of patients with eso-phagus or esophagogastric cancer were included.HSP-positive patients were compared with HSP-negative,and the endpoints analyzed were lymph node metastasis,tumor depth,distant metastasis,and overall survival(OS).HSPs were stratified according to the HSP family,and the summary risk difference(RD)was calculated using a random-effect model.RESULTS The final selection comprised 27 studies,including esophageal squamous cell carcinoma(21),esophagogastric adenocarcinoma(5),and mixed neoplasms(1).The pooled sample size was 3465 patients.HSP40 and 60 were associated with a higher 3-year OS[HSP40:RD=0.22;95%confidence interval(CI):0.09-0.35;HSP60:RD=0.33;95%CI:0.17-0.50],while HSF1 was associated with a poor 3-year OS(RD=-0.22;95%CI:-0.32 to-0.12).The other HSP families were not associated with long-term survival.HSF1 was associated with a higher probability of lymph node metastasis(RD=-0.16;95%CI:-0.29 to-0.04).HSP40 was associated with a lower probability of lymph node dissemination(RD=0.18;95%CI:0.03-0.33).The expression of other HSP families was not significantly related to tumor depth and lymph node or distant metastasis.CONCLUSION The expression levels of certain families of HSP,such as HSP40 and 60 and HSF1,are associated with long-term survival and lymph node dissemination in patients with esophageal and esophagogastric cancer.

Characterization of physicochemical and immunogenic properties of allergenic proteins altered by food processing:a review

Food allergens are mainly naturally-occurring proteins with immunoglobulin E(IgE)-binding epitopes.Understanding the structural and immunogenic characteristics of allergenic proteins is essential in assessing whether and how food processing techniques reduce allergenicity.We here discuss the impacts of food processing technologies on the modification of physicochemical,structural,and immunogenic properties of allergenic proteins.Detection techniques for characterizing changes in these properties of food allergens are summarized.Food processing helps to reduce allergenicity by aggregating or denaturing proteins,which masks,modifies,or destroys antigenic epitopes,whereas,it cannot eliminate allergenicity completely,and sometimes even improves allergenicity by exposing new epitopes.Moreover,most food processing techniques have been tested on purified food allergens rather than food products due to potential interference of other food components.We provide guidance for further development of processing operations that can decrease the allergenicity of allergenic food proteins without negatively impacting the nutritional profile.

Interplay between the glymphatic system and neurotoxic proteins in Parkinson’s disease and related disorders:current knowledge and future directions

Parkinson’s disease is a common neurodegenerative disorder that is associated with abnormal aggregation and accumulation of neurotoxic proteins,includingα-synuclein,amyloid-β,and tau,in addition to the impaired elimination of these neurotoxic protein.Atypical parkinsonism,which has the same clinical presentation and neuropathology as Parkinson’s disease,expands the disease landscape within the continuum of Parkinson’s disease and related disorders.The glymphatic system is a waste clearance system in the brain,which is responsible for eliminating the neurotoxic proteins from the interstitial fluid.Impairment of the glymphatic system has been proposed as a significant contributor to the development and progression of neurodegenerative disease,as it exacerbates the aggregation of neurotoxic proteins and deteriorates neuronal damage.Therefore,impairment of the glymphatic system could be considered as the final common pathway to neurodegeneration.Previous evidence has provided initial insights into the potential effect of the impaired glymphatic system on Parkinson’s disease and related disorders;however,many unanswered questions remain.This review aims to provide a comprehensive summary of the growing literature on the glymphatic system in Parkinson’s disease and related disorders.The focus of this review is on identifying the manifestations and mechanisms of interplay between the glymphatic system and neurotoxic proteins,including loss of polarization of aquaporin-4 in astrocytic endfeet,sleep and circadian rhythms,neuroinflammation,astrogliosis,and gliosis.This review further delves into the underlying pathophysiology of the glymphatic system in Parkinson’s disease and related disorders,and the potential implications of targeting the glymphatic system as a novel and promising therapeutic strategy.

CRISPR/Cas9-mediated NlInR2 mutants:Analyses of residual mRNA and truncated proteins

CRISPR/Cas9 technology is a powerful genome manipulation tool in insects.However,little is known about whether mRNA and protein of a target gene are completely cleared in homozygous mutants.This study generated homozygous mutants of the insulin receptor gene 2(NlInR2)in the brown planthopper(Nilaparvata lugens)using CRISPR/Cas9 genome editing.Both frameshift mutants,E5_D17 and E6_I7,differentiated towards long wings,but there were differences in wing morphology,with E5_D17 showing wing deformities.Subsequent investigations revealed the presence of residual expression of NlInR2 mRNA in both mutants,as well as the occurrence of spliceosomes featuring exon skipping splicing in E5_D17.Additionally,the E5_D17 exhibited the detection of N-terminally truncated NlInR2 protein.RNA interference experiments indicated that the knockdown of NlInR2 expression in the E5_D17 mutant line increased the proportion of wing deformities from 11.1 to 65.6%,suggesting that the residual NlInR2 mRNA of the E5_D17 mutant might have retained some genetic functions.Our results imply that systematic characterization of residual protein expression or function in CRISPR/Cas9-generated mutant lines is necessary for phenotypic interpretation.

Exploration of cyclooxygenase-2 inhibitory peptides from walnut dreg proteins based on in silico and in vitro analysis

Walnut dreg protein hydrolysates(WDPHs)exhibit a variety of biological activities,however,the cyclooxygenase-2(COX-2)inhibitory peptide of WDPHs remain unclear.The aim of this study was to rapidly screen for such peptides in WDPHs through a combination of in silico and in vitro analysis.In total,1262 peptide sequences were observed by nano liquid chromatography/tandem mass spectrometry(nano LC-MS/MS)and 4 novel COX-2 inhibitory peptides(AGFP,FPGA,LFPD,and VGFP)were identified.Enzyme kinetic data indicated that AGFP,FPGA,and LFPD displayed mixed-type COX-2 inhibition,whereas VGFP was a non-competitive inhibitor.This is mainly because the peptides form hydrogen bonds and hydrophobic interactions with residues in the COX-2 active site.These results demonstrate that computer analysis combined with in vitro evaluation allows for rapid screening of COX-2 inhibitory peptides in walnut protein dregs.

Analysis and Review of Downregulated Actin Cytoskeletal Proteins in Non-Small Cell Lung Cancer

Actin, a highly conserved protein, plays a dominant role in Non-small cell lung cancer (NSCLC). Late diagnosis and the aggressive nature of NSCLC pose a significant threat. Studying the clinic pathological properties of NSCLC proteins is a potential alternative for developing treatment strategies. Towards this, 35 downregulated actin cytoskeletal proteins on NSCLC prognosis and treatment were studied by examining their protein-protein interactions, gene ontology enrichment terms, and signaling pathways. Using PubMed, various proteins in NSCLC were identified. The protein-protein interactions and functional associations of these proteins were examined using the STRING database. The focal adhesion signaling pathway was selected from all available KEGG and Wiki pathways because of its role in regulating gene expression, facilitating cell movement and reproduction, and significantly impacting NSCLC. The protein-protein interaction network of the 35 downregulated actin cytoskeleton proteins revealed that ACTG1, ACTR2, ACTR3, ANXA2, ARPC4, FLNA, TLN1, CALD1, MYL6, MYH9, MYH10, TPM1, TPM3, TPM4, PFN1, IQGAP1, MSN, and ZXY exhibited the highest number of interactions. Whereas HSPB1, CTNNA1, KRT17, KRT7, FLNB, SEPT2, and TUBA1B displayed medium interactions, while UTRN, TUBA1B, and DUSP23 had relatively fewer interactions. It was discovered that focal adhesions are critical in connecting membrane receptors with the actin cytoskeleton. In addition, protein kinases, phosphatases, and adapter proteins were identified as key signaling molecules in this process, greatly influencing cell shape, motility, and gene expression. Our analysis shows that the focal adhesion pathway plays a crucial role in NSCLC and is essential for developing effective treatment strategies and improving patient outcomes.

Research Progress on Plant Anti-Freeze Proteins

Plant antifreeze proteins(AFPs)are special proteins that can protect plant cells from ice crystal damage in low-temperature environments,and they play a crucial role in the process of plants adapting to cold environ-ments.Proteins with these characteristics have been found infish living in cold regions,as well as many plants and insects.Although research on plant AFPs started relatively late,their application prospects are broad,leading to the attention of many researchers to the isolation,cloning,and genetic improvement of plant AFP genes.Studies have found that the distribution of AFPs in different species seems to be the result of independent evolu-tionary events.Unlike the AFPs found infish and insects,plant AFPs have multiple hydrophilic ice-binding domains,and their recrystallization inhibition activity is about 10–100 times that offish and insect AFPs.Although different plant AFPs have the characteristics of low TH and high RI,their DNA and amino acid sequences are completely different,with small homology.With in-depth research and analysis of the character-istics and mechanisms of plant AFPs,not only has our understanding of plant antifreeze mechanisms been enriched,but it can also be used to improve crop varieties and enhance their freezing tolerance,yield,and quality through genetic engineering.In addition,the study of plant AFPs also contributes to our understanding of freezing resistance mechanisms in other organisms and provides new research directions for thefield of biotech-nology.Therefore,based on the analysis of relevant literature,this article will delve into the concepts,character-istics,research methods,and mechanisms of plant AFPs,summarize the latest research progress and application prospects of AFPs in plant,and provide prospects for the future development of AFP gene research.

Changes in milk fat globule membrane proteins along lactation stage of Laoshan dairy goat

The milk fat globule membrane(MFGM)is a complex structure with numerous functions,and its composition is affected by many factors.There have been few systematic investigations on goat MFGM proteome profiling during lactation.Individual milk samples from 15 healthy dairy goats were obtained at six lactation time points for investigation of the MFGM proteome using both data-independent acquisition(DIA)and data-dependent acquisition(DDA)proteomics techniques combined with multivariate statistical analysis.Using the DIA method,890 variably abundant MFGM proteins were discovered throughout the lactation cycle.From 1 to 240 d,butyrophilin subfamily 1 member A1,lipoprotein lipase,perilipin-2,and adipose triglyceride lipase were upregulated,while APOE,complement C3,clusterin,and IgG were downregulated.Furthermore,from 1 to 90 d,annexin A1,annexin A2,and antithrombin-ll were downregulated,then upregulated by d 240.Albumin had a high degree of connectedness,indicating that it was a key protein,according to protein-protein interaction research.Overall,our findings gave new insights into the biological features of MFGM protein in goat milk throughout lactation,which may aid in the creation of specialized MFGM products and infant formula.

Hmo1:A versatile member of the high mobility group box family of chromosomal architecture proteins

Eukaryotic chromatin consisting of nucleosomes connected by linker DNA is organized into higher order structures,which is facilitated by linker histone H1.Formation of chromatin compacts and protects the genome,but also hinders DNA transactions.Cells have evolved mechanisms to modify/remodel chromatin resulting in chromatin states suitable for genome functions.The high mobility group box(HMGB)proteins are non-histone chromatin architectural factors characterized by one or more HMGB motifs that bind DNA in a sequence nonspecific fashion.They play a major role in chromatin dynamics.The Saccharomyces cerevisiae(yeast hereafter)HMGB protein Hmo1 contains two HMGB motifs.However,unlike a canonical HMGB protein that has an acidic C-terminus,Hmo1 ends with a lysine rich,basic,C-terminus,resembling linker histone H1.Hmo1 exhibits characteristics of both HMGB proteins and linker histones in its multiple functions.For instance,Hmo1 promotes transcription by RNA polymerases I and II like canonical HMGB proteins but makes chromatin more compact/stable like linker histones.Recent studies have demonstrated that Hmo1 destabilizes/disrupts nucleosome similarly as other HMGB proteins in vitro and acts to maintain a common topological architecture of genes in yeast genome.This minireview reviews the functions of Hmo1 and the underlying mechanisms,highlighting recent discoveries.

Analysis of quality-related proteins in golden pompano(Trachinotus ovatus)fillets with modified atmosphere packaging under superchilling storage

Here,we aimed to study the changes in proteome of golden pompano fillets during post-mortem storage.Tandem mass tags(TMT)-labeled quantitative proteomic strategy was applied to investigate the relationships between protein changes and quality characteristics of modified atmosphere packaging(MAP)fillets during superchilling(-3°C)storage.Scanning electron microscopy was used to show that the muscle histology microstructure of fillets was damaged to varying degrees,and low-field nuclear magnetic resonance was used to find that the immobilized water and free water in the muscle of fillets changed significantly.Total sulfhydryl content,TCA-soluble peptides and Ca2+-ATPase activity also showed that the fillet protein had a deterioration by oxidation and denaturation.The Fresh(FS),MAP,and air packaging(AP)groups were set.Total of 150 proteins were identified as differential abundant proteins(DAPs)in MAP/FS,while 209 DAPs were in AP/FS group.The KEGG pathway analysis indicated that most DAPs were involved in binding proteins and protein turnover.Correlation analysis found that 52 DAPs were correlated with quality traits.Among them,8 highly correlated DAPs are expected to be used as potential quality markers for protein oxidation and water-holding capacity.These results provide a further understanding of the muscle deterioration mechanism of packaging golden pompano fillets during superchilling.

The Relationship Between Developmental Accumulation of Leaf Soluble Proteins and Vernalization Response of Wheat(Triticum aestivum L.em. Thell)

The relationship between vernalization requirement and quantitative and qualitative changes in total leaf soluble proteins were determined in one spring （cv. Kohdasht） and two winter （cvs. Sardari and Norstar） cultivars of wheat （Triticum aestivum L.） exposed to 4℃. Plants were sampled on days 2, 14, 21 and 35 of exposure to 4℃. The final leaf number （FLN） was determined throughout the vernalization periods （0, 7, 14, 24, and 35 d） at 4℃. The final leaf number decreased until days 24 and 35 in Sardari and Norstar eultivars, respectively, indicating the vernalization saturation at these times. No clear changes were detected in the final leaf number of Kohdash cultivar, verifying no vernalization requirement for this spring wheat cultivar. Comparing with control, clear cold-induced 2-fold increases in proteins quantity occurred after 48 h following the 4℃-treatment in the leaves of the both winter wheat cultivars but, such response was not detected in the spring cultivar. However, the electrophoretic protein patterns showed between-cultivar and between-temperature treatment differences. With increasing exposure time to 4℃, the winter cultivars tended to produce more HMW polypeptides than the spring cultivar. Similar proteins were induced in both Sardari and Norstar winter wheat cultivars, however, the long vernalization requirement in Norstar resulted in high level and longer duration of expression of cold-induced proteins compared to Sardari with a short vernalization requirement. These observations indicate that vernalization response regulates the expression of low temperature （LT） tolerance proteins and determines the duration of expression of LT- induced proteins.

Soluble expression of recomb inant cMyc,Klf4,Oct4,and Sox2 proteins in bacteria and transduction into living cells

AIM：To develop a new method to produce recombinant reprogramming proteins,c Myc,Klf4,Oct4,and Sox2,in soluble format with low cost for the generation of induced pluripotent stem cells（i PSCs）.METHODS：A short polypeptide sequence derived from the HIV trans-activator of transcription protein（TAT） and the nucleus localization signal（NLS） polypeptide were fused to the N terminus of the reprogramming proteins and they were constructed into p Cold-SUMO vector which can extremely improve the solubility of recombinant proteins.Then these vector plasmids were transformed into E.coli BL21（DE3） Chaperone competent cells for amplification.The solubility of these recombinant proteins was determined by SDS-PAGE and Coomassie brilliant blue staining.The recombinant proteins were purified by NiNTA resin and identified by Western blot.The transduction of these proteins into HEK 293 T cells were evaluated by immunofluorescence staining.RESULTS：These four reprogramming proteins could be produced in soluble format in p Cold-SUMO expression vector system with the assistance of chaperone proteins in bacteria.The proteins were purified successfully with a purity of over 70% with a relative high transduction rate into 293 cells.CONCLUSION：The results in the present study indicate the four important reprogramming proteins,c Myc,Klf4,Oct4,and Sox2,can be produced in soluble format in bacteria with low cost.Our new method thus might be expected to greatly contribute to the future study of i PSCs.

Identification,evolution,expression and protein interaction analysis of genes encoding B-box zinc-finger proteins in maize

The B-box(BBX)family of proteins consists of zinc-finger transcription factors with one or two highly conserved B-box motifs at their N-termini.BBX proteins play crucial roles in various aspects of plant growth and development,including seedling photomorphogenesis,shade avoidance,flowering time,and biotic and abiotic stress responses.Previous studies have identified many different BBXs from several plant species,although the BBX family members in maize are largely unknown.Genome-wide identification and comprehensive analysis of maize BBX(ZmBBX)expression and interaction networks would therefore provide valuable information for understanding their functions.In this study,36 maize BBXs in three major clades were identified.The ZmBBXs within a given clade were found to share similar domains,motifs,and genomic structures.Gene duplication analyses revealed that the expansion of BBX proteins in maize has mainly occurred by segmental duplication.The expression levels of ZmBBXs were analyzed in various organs and tissues,and under different abiotic stress conditions.Protein–protein interaction networks of ZmBBXs were established using bioinformatic tools and verified by bimolecular fluorescence complementation(BiFC)assays.Our findings can facilitate a greater understanding of the complexity of the ZmBBX family and provide novel clues for unravelling ZmBBX protein functions.

Soluble Structure of CLIC and S100 Proteins Investigated by Atomic Force Microscopy

The ability to visualise proteins in their native environment and discern information regarding stoichiometry is of critical importance when studying protein interactions and function. We have used liquid cell atomic force microscopy (AFM) to visualise proteins in their native state in buffer and have determined their molecular volumes. The human proteins S100A8, S100A9, S100A12 and CLIC1 were used in this investigation. The effect of oxidation on the protein structure of CLIC1 was also investigated and we found that CLIC1 multimerisation could be discerned by AFM, which supports similar findings by other methods. We have found good correlation between the molecular volumes measured by AFM and the calculated volumes of the individual proteins. This method allows for the study of single soluble proteins under physiological conditions and could potentially be extended to study the structure of these proteins when located within a membrane environment.

Rice Storage Proteins:Focus on Composition,Distribution,Genetic Improvement and Effects on Rice Quality

Rice storage proteins(RSPs)are plant proteins with high nutritional quality.As the second largest type of storage substance in rice,it is the main source of protein intake for people who consume rice as a staple food.The content and type of RSPs affect the appearance,processing quality and eating quality of rice.These effects involve the distribution of RSPs in rice grains as well as the interactions of RSPs with other components such as starch in rice grains.In the past two decades,some progress has been made in the genetic improvement of RSPs.However,the determination mechanism of protein content and composition in rice is still unclear,and the mechanism of the effect of RSPs on rice quality has not been elucidated.In this review,the composition,biosynthesis and distribution of RSPs,and quantitative trait loci mapping and cloning of RSP genes are summarized,the research progress of the influence of RSPs and their components on rice quality are reviewed,and the research directions in the future are proposed.

Favorable and unfavorable amino acid residues in water-soluble and transmembrane proteins

We analyzed the amino acid residues present in the water-soluble and transmembrane proteins of 6 thermophilic and 6 mesophilic species of the domains Archaea and Eubacteria, and characterized them as favorable or unfavorable. The characterization was performed by comparing the observed number of each amino acid residue to the expected number calculated from the percentage of nucleotides present in each gene. Amino acids that were more or less abundant than expected were considered as favorable or unfavorable, respectively. Comparisons of amino acid compositions indicated that the water-soluble proteins were rich in charged residues such as Glu, Asp, Lys, and His, whereas hydrophobic residues such as Trp, Phe, and Leu were abundant in transmembrane proteins. Interestingly, our results found that although the Trp residue was abundant in transmembrane proteins, it was not defined as favorable by our calculations, indicating that increased numbers of a particular amino acid does not necessary indicate it is a favorable residue. Amino acids with high G + C content such as Ala, Gly, and Pro were frequently observed as favorable in species with low G + C content. Comparatively, amino acids with low G + C content such as Phe, Tyr, Lys, Ile, and Met were frequently observed as favorable in species with high G + C content. These are the examples to increase the supply of amino acids than expected. Amino acids with neutral G + C content, i.e., Glu and Asp were favorable in water-soluble proteins from all species analyzed, and Cys was unfavorable both in water-soluble and transmembrane proteins. These results indicate that amino acid compositions are essentially determined by the nucleotide sequence of the genes, and the amino acid content is altered by a deviation from expectation.

Histone H3K27me3 methylation regulates the expression of secreted proteins distributed at fast-evolving regions through transcriptional repression of transposable elements

The fine-tuned expression dynamics of the effector genes are pivotal for the transition from vegetative growth to host colonization of pathogenic filamentous fungi.However,mechanisms underlying the dynamic regulation of these genes remain largely unknown.Here,through comparative transcriptome and chromatin immunoprecipitation sequencing(ChIP-seq)analyses of the methyltransferase PoKmt6 in rice blast fungus Pyricularia oryzae(syn.Magnaporthe oryzae),we found that PoKmt6-mediated H3K27me3 deposition was enriched mainly at fast-evolving regions and contributed to the silencing of a subset of secreted proteins(SP)and transposable element(TE)families during the vegetative growth of P.oryzae.Intriguingly,we observed that a group of SP genes,which were depleted of H3K27me3 modification,could also be silenced via the H3K27me3-mediated repression of the nearby TEs.In conclusion,our results indicate that H3K27me3 modification mediated by PoKmt6 regulates the expression of some SP genes in fast-evolving regions through the suppression of nearby TEs.

Eukaryotic expression, purification and activity characterization of human soluble DSG2 extracellular domain protein

Objective:To construct a secretory eukaryotic expression vector of DSG2 fused with the Fc region of the human IgG,to validate its expression in 293T cells,and to purify the secretory protein with biological activity.Methods:The DSG2 extracellular domain fragment gene(DSG2ex),was amplified by PCR,and was inserted into the eukaryotic expression plasmid pCMV3-IgG1 to construct the recombinant eukaryotic expression plasmid-pCMV3-DSG2ex-IgG1.The successfully constructed eukaryotic expression plasmid was transfected into 293T cells to express and secrete DSG2 extracellular domain protein.The targeted protein was purified from the cell culture supernatant by Protein A affinity chromatography and confirmed by Western Blotting and ELISA.Results:The pCMV3-DSG2ex-IgG1 eukaryotic expression plasmid was successfully constructed.The highest protein expression level was obtained with 293T cells after 96 h of transfection.The relative molecular mass of the purified product was between 100 and 130 kDa was estimated by SDS-PAGE,which was consistent with the expectation.The yield of the purified protein reached 0.8 mg/ml with a purity over 90%.The purified DSG2 extracellular domain protein with IgG1 tag was recognized by IgG monoclonal antibodies by Western blotting.Moreover,the ELISA results showed that the prepared DSG2 extracellular domain protein had significant binding activity to human type 55 adenovirus Fiber Knob protein(HAdV-55).Conclusion:A simple and efficient method for eukaryotic expression and purification of human soluble DSG2 extracellular domain protein was successfully established,and biologically active DSG2 extracellular domain protein was purified,which laid the foundation for the later study of its protein function and anti-adenovirus drugs.