Search

Abstract This study aimed to analyze anthropometric and physiological profiles of highly trained sailors and the differences between sailors regarding various training levels. Forty-two sailors (22 male, 22.4 ± 3.8 years; 20 females, 21.3 ± 3.6 years) were divided into helmsmen and crew groups, and the high- and low-level were distinguished. Sailors completed height, sitting height, legs length, weight, BMI, VO2max, 30 s all-out sprint, isometric mid-thigh pull (IMTP), countermovement jump, bench pull, core endurance tests. The results showed the crew had higher height, sitting height, weight, VO2max and lower trunk flexor endurance test times compared to the helmsmen (p

ObjectivesThe role of a coach in enhancing athletes’ performance and achieving success is well-documented across numerous studies. However, the strategies employed by Chinese coaches in developing world champion gymnasts remain under explored.MethodsThis research involved a single case study focusing on a coach from the Chinese National Men’s Gymnastics Team, credited with nurturing eight world champion gymnasts.ResultsThe inductive content analysis leads to that 6 subthemes, “international perspective and collaborative ability,” “ability to control and regulate training loads,” “identifying athletes’ needs and transforming them into motivation,” “goal setting aligned with athletes’ abilities,” “adopting authoritative democratic coaching style,” and “establishing hierarchical-style friendship”, and 3 themes, “training management and planning,” “motivation and goal setting,” and “interpersonal communication” are manifested. An overarching theme “the successful experience of gymnastics world champion coach,” is derived from the analysis.ConclusionThis research bridges the gap between theoretical knowledge and practical application, offering valuable insights into the successful experiences of gymnastics world champion coaches. The findings have the potential to influence coaching methodologies globally, fostering the development of resilient, motivated, and high-performing athletes. Future research should focus on sport-specific studies, longitudinal analyses, and cross-cultural comparisons to further advance the field of sports coaching and validate the effectiveness of these innovative coaching strategies.

ObjectivesThis study explores the significant impact of expressive ties (EI) between DanceSport couples on their competitive performance (CP). Utilizing a dyadic approach, we examined the performance achievement processes of DanceSport couples in relation to their EI.MethodsParticipants comprised 67 dyads of Chinese elite dancers aged between 16 and 30 years. The dyadic analysis was carried out using a structural equation model based on the actor-partner interdependence mediation model.ResultsWith regard to actor effects, both male (β = 0.292, p = 0.012) and female (β = 0.443, p

Objective: To assess the effects of 8 weeks of unilateral (UNI), bilateral (BI), and combined unilateral + bilateral (UNI + BI) resistance training on bench press and squat strength in adolescent boxers.Methods: Using the Gym Aware linear accelerometer, free-weight squat and bench press strength exercises were evaluated after an 8-week training intervention. Thirty adolescent boxers were randomly assigned to three groups: UNI, height: 1.73 ± 0.08 m, weight: 55.42 ± 5.85 kg; UNI + BI, height: 1.7 ± 0.06 m, weight: 54.73 ± 5.33 kg; and BI, height: 1.74 ± 0.06 m, weight: 59.67 ± 8.39 kg. Each group followed their designated UNI/BI/UNI + BI compound resistance training protocols, and the effects of 8 weeks of single-sided and bilateral intervention training on the performance of free-weight squat and bench press exercises at 30%, 50%, and 80% of 1-repetition maximum (1RM) were evaluated.Results: Significant improvements were observed in the 30% 1RM, 50% 1RM, and 80% 1RM outcomes for both squat and bench press exercises before and after the interventions (p < 0.05, p < 0.01). In the intergroup comparison, GymAware measurements revealed that the UNI and UNI + BI groups exhibited superior peak power values for squat and bench press exercises at 30% 1RM compared to the BI group.Discussion: UNI and UNI + BI training led to significantly higher output power values in bench press and squat exercises at 30% 1RM compared to the BI training group.

Abstract In the post-genomic era, the demand for faster and more efficient protein production has increased, both in public laboratories and industry. In addition, with the expansion of protein sequences in databases, the range of possible enzymes of interest for a given application is also increasing. Faced with peer competition, budgetary, and time constraints, companies and laboratories must find ways to develop a robust manufacturing process for recombinant protein production. In this review, we explore high-throughput technologies for recombinant protein expression and present a holistic high-throughput process development strategy that spans from genes to proteins. We discuss the challenges that come with this task, the limitations of previous studies, and future research directions.

Abstract All-trans-retinoic acid (ATRA), the retinoic acid receptors (RARs) agonist, regulates cell growth, differentiation, immunity, and survival. We report that ATRA-treatment repressed cancer growth in syngeneic immunocompetent, but not immunodeficient mice. The tumor microenvironment was implicated: CD8+ T cell depletion antagonized ATRA’s anti-tumorigenic effects in syngeneic mice. ATRA-treatment with checkpoint blockade did not cooperatively inhibit murine lung cancer growth. To augment ATRA’s anti-tumorigenicity without promoting its pro-tumorigenic potential, an RARγ agonist (IRX4647) was used since it regulates T cell biology. Treating with IRX4647 in combination with an immune checkpoint (anti-PD-L1) inhibitor resulted in a statistically significant suppression of syngeneic 344SQ lung cancers in mice—a model known for its resistance to checkpoints and characterized by low basal T cell and PD-L1 expression. This combined treatment notably elevated CD4+ T-cell presence within the tumor microenvironment and increased IL-5 and IL-13 tumor levels, while simultaneously decreasing CD38 in the tumor stroma. IL-5 and/or IL-13 treatments increased CD4+ more than CD8+ T-cells in mice. IRX4647-treatment did not appreciably affect in vitro lung cancer growth, despite RARγ expression. Pharmacokinetic analysis found IRX4647 plasma half-life was 6 h in mice. Yet, RARα antagonist (IRX6696)-treatment with anti-PD-L1 did not repress syngeneic lung cancer growth. Together, these findings provide a rationale for a clinical trial investigating an RARγ agonist to augment check point blockade response in cancers.

ObjectiveSmartphone addiction, as a key topic in the current field of behavioral addictions and public health, has brought many negative impacts on the physical, psychological, interpersonal communication, and even academic performance among contemporary college students. Therefore, the purpose of this study is to offer ideas for solving smartphone addiction among college students through investigating the potential mediating effect of self-esteem in the relationship between physical activity and smartphone addiction.MethodsBy the quota sampling, a cross-sectional study was conducted to investigate 650 college students from 10 colleges in Guangzhou Higher Mega Center, and several self-reported instruments including physical activity rating scale-3 (PARS-3), mobile phone addiction tendency scale (MPATS), self-esteem scale (SES) were used to collect the related data needed for the present study. The descriptive analysis, correlation analysis, hierarchical regression analysis, and mediating effect analysis in this study were performed in turn.ResultsThe results showed that physical activity (r = −0.124, p < 0.01) and self-esteem (r = −0.360, p < 0.01) were all negatively correlated with smartphone addiction, and both could also significantly and negatively predict smartphone addiction. There was a positive correlation between physical activity and self-esteem (r = 0.084, p < 0.05), and self-esteem could be significantly predicted by physical activity. And more important, the relationship between physical activity and smartphone addiction could be partially mediated by self-esteem, and the indirect effect value was −0.346 (95% Boot CI = −0.695; −0.023), along with the mediating effect accounted for 24% of the total effect between physical activity and smartphone addiction.ConclusionThe current study shows that physical activity could not only directly reduce smartphone addiction, but also decrease smartphone addiction by indirectly improving self-esteem, which is important in practice for solving this troublesome issue and then gradually developing a healthy behavior in daily life for college students in China, and even across the world in near future.

ObjectivesAlthough the positive association of partnership between DanceSport couples with competitive performance (CP) is documented, less is known about the mediating factors of this relationship. According to the related literature and self-determination theory (SDT), the present study finds and verifies that athlete engagement (AE) mediates the association between partnership and competitive performance.MethodsA total of 242 Chinese sports dancers were recruited using the purposive sampling method. The Partnership Scale-DanceSport Couples (PS-DSC), the Athlete Engagement Questionnaire (AEQ), and the Competitive Performance Questionnaire (CPQ) were adopted to collect data.ResultsThe obligatory instrumental ties, expressive ties, and interpersonal perception scores are all positively correlated with both athlete engagement and competitive performance, and athlete engagement scores are positively correlated with competitive performance. Athlete engagement completely mediates the association between obligatory instrumental ties and competitive performance, and it partially mediates the association between expressive ties, interpersonal perception, and competitive performance, with the mediating effect accounting for 25.29 and 24.40% of the total effect, respectively.ConclusionAthlete engagement mediates the association between DanceSport couples’ partnership and competitive performance. High levels of athlete engagement are needed to improve the chance of promoting obligatory instrumental ties, expressive ties, and interpersonal perception between DanceSport couples toward excellent competitive performance. Overall, the results represent an attempt to extend our understanding of the mechanisms by which the three partnership stereotype factors individually influence dancers’ cognitive and psychological states.

Apple ring rot, which is caused by Botryosphaeria dothidea, severely affects apple production. The mechanisms employed in apple cells against B. dothidea remain unknown. In this research, the pathogen infection mode and the relationship between cell death and disease resistance in ‘Fuji’/B. dothidea interaction pathosystem were investigated. By using transmission electron microscopy (TEM), our research showed that the pathogen infects apple cells both intracellularly and extracellularly. However, compared with that in immature fruit, the incidence of hyphae in the interior of mature apple fruit cells increased dramatically, suggesting that cell wall-mediated penetration resistance could be important in apple resistance against B. dothidea. TEM ultrastructural characterization identified the nuclear morphology of programmed cell death induction in both apple fruit and callus cells under B. dothidea infection. Overexpression of MdVDAC2 (MDP0000271281), which encodes an outer-membrane localized anion channel protein in mitochondria, significantly promoted cell death under B. dothidea infection and simultaneously inhibited pathogen infection, suggesting that cell death represents a disease resistance mechanism in apple against B. dothidea infection. Furthermore, BdCatalase (KAF4307763), a cytochrome P450 family protein BdCYP52A4 (KAF4300696), and subtilisin-domain containing proteins were identified from B. dothidea-secreted proteins, which suggested the potential involvement of active oxygen species and phytoalexins in combating B. dothidea infection and triggering or dampening apple resistance. Collectively, our research suggested that cell wall-mediated penetration resistance, programmed cell death machinery and microbial effector-interrelated signaling were among strategies recruited in apple to combat B. dothidea. The current research laid the foundation for further investigations into resistance mechanisms in apple.

Cancer metastasis is a major cause of cancer-related mortality. Strategies to reduce metastases are needed especially in lung cancer, the most common cause of cancer mortality. We previously reported increased ubiquitin-specific peptidase 18 (USP18) expression in lung and other cancers. Engineered reduction of USP18 expression repressed lung cancer growth and promoted apoptosis. This deubiquitinase (DUB) stabilized targeted proteins by removing the complex interferon-stimulated gene 15 (ISG15). This study explores if the loss of USP18 reduced lung cancer metastasis. USP18 knock-down in lung cancer cells was independently achieved using small hairpin RNAs (shRNAs) and small interfering RNAs (siRNAs). USP18 knock-down reduced lung cancer growth, wound-healing, migration, and invasion versus controls (P

Corynebacterium glutamicum represents an emerging recombinant protein expression factory due to its ideal features for protein secretion, but its applicability is harmed by the lack of an autoinduction system with tight regulation and high yield. Here, we propose a new recombinant protein manufacturing platform that leverages ethanol as both a delayed carbon source and an inducer. First, we reanalysed the native inducible promoter PICL from the acetate uptake operon and found that its limited capacity is the result of the inadequate translation initial architecture. The two strategies of bicistronic design and ribozyme-based insulator can ensure the high activity of this promoter. Next, through transcriptional engineering that alters transcription factor binding sites (TFBSs) and the first transcribed sequence, the truncated promoter PA256 with a dramatically higher transcription level was generated. When producing the superfolder green fluorescent protein (sfGFP) under 1% ethanol conditions, PA256 exhibited substantially lower protein accumulation in prophase but an approximately 2.5-fold greater final yield than the strong promoter PH36. This superior expression mode was further validated using two secreted proteins, camelid antibody fragment (VHH) and endoxylanase (XynA). Furthermore, utilizing CRISPRi technology, ethanol utilization blocking strains were created, and PA256 was shown to be impaired in the phosphotransacetylase (PTA) knockdown strains, indicating that ethanol metabolism into the tricarboxylic acid cycle is required for PA256 upregulation. Finally, this platform was applied to produce the “de novo design” protein NEO-2/15, and by introducing the N-propeptide of CspB, NEO-2/15 was effectively secreted with the accumulation 281 mg/L obtained after 24 h of shake-flask fermentation. To the best of our knowledge, this is the first report of NEO-2/15 secretory overexpression.

IntroductionDanceSport is described as a dance involving a male–female partner. It is important to comprehend the partnership between dance couples so that their competitive performance can be effectively supported. However, only a few studies have verified the influence of partnership between DanceSport couples on competitive performance and explored its psychological mechanism to provide means to deal with the partnership. The reason was that there was a lack of appropriate assessment tools.AimsThis multi-study outlines the development, content, and construct validity of a novel, mixed-method tool to assess DanceSport partnership.MethodsThe development of the Partnership Scale-DanceSport Couples (PS-DSC) included four studies and data from four samples of Chinese elite dancers (N = 914 total). In stage 1, outlined in study 1, PS-DSC items were generated and then refined using the feedback provided by academics, sports coaches, and elite dancers. In stage 2, outlined in studies 2 and 3, exploratory factor analysis and confirmatory factor analysis were used to examine the structure of the PS-DSC items. In stage 3, outlined in study 4, composite reliability, discriminant validity, and convergent validity were assessed. The result of this process was a 13-item three-factor instrument. Based on these initial findings, the PS-DSC provided the first valid and reliable way of measuring partnerships between DanceSport couples.ConclusionThis study has taken the promising first step in developing a tool to comprehensively measure partnerships between DanceSport couples.

Linoleic acid (LA), a potentially algae-inhibiting chemical released by macroalgae, has been shown to hinder the growth of numerous bloom-forming species. The allelopathic effects of LA (varying from 100 μg/L to 900 μg/L) on harmful microalgae K. mikimotoi were examined using population growth dynamics and physiological levels of K. mikimotoi. LA (>500 μg/L) strongly inhibited algal growth with most cells halted at the S and G2 phases and an evident drop in photosynthetic pigments (chlorophyll a (chl a), chlorophyll c (chl c) and carotenoids). Furthermore, chlorophyll fluorescence parameters such as Fv/Fm, PI, ETo/RC showed a declining trend whereas ABS/RC, DIo/RC, TRo/RC showed an increasing trend with increasing LA exposure concentrations. The level of intracellular reactive oxygen species (ROS) was considerably higher, indicating that LA promoted oxidative stress in K. mikimotoi. Excessive ROS promoted apoptosis in K. mikimotoi, which was noted by increased activity of caspase-3, caspase-9, and flow cytometry (FCM) data. Furthermore, N-acetylcysteine (NAC) and N-Acetyl-Asp-Glu-Val-Asp-CHO (Ac-DEVD-CHO) lowered the apoptotic rates of the LA-treated algal cells, indicating that the aforementioned inhibitors delayed K. mikimotoi apoptosis under LA treatment. To summarize, cell cycle arrest of K. mikimotoi is less sensitive to ROS, but the overproduction of ROS generated by LA activated caspase-3 and caspase-9, which further promoted the apoptosis of K. mikimotoi. This research showed that LA might have great potential and application prospects in controlling the outbreak of harmful algae.

Abstract Terpenoids form the most diversified class of natural products, which have gained application in the pharmaceutical, food, transportation, and fine and bulk chemical industries. Extraction from naturally occurring sources does not meet industrial demands, whereas chemical synthesis is often associated with poor enantio-selectivity, harsh working conditions, and environmental pollutions. Microbial cell factories come as a suitable replacement. However, designing efficient microbial platforms for isoprenoid synthesis is often a challenging task. This has to do with the cytotoxic effects of pathway intermediates and some end products, instability of expressed pathways, as well as high enzyme promiscuity. Also, the low enzymatic activity of some terpene synthases and prenyltransferases, and the lack of an efficient throughput system to screen improved high-performing strains are bottlenecks in strain development. Metabolic engineering and synthetic biology seek to overcome these issues through the provision of effective synthetic tools. This review sought to provide an in-depth description of novel strategies for improving cell factory performance. We focused on improving transcriptional and translational efficiencies through static and dynamic regulatory elements, enzyme engineering and high-throughput screening strategies, cellular function enhancement through chromosomal integration, metabolite tolerance, and modularization of pathways. Graphical Abstract

The title compound, C19H13N, which crystallizes in the monoclinic C2/c space group with one half-molecule in the asymmetric unit, was synthesized by Suzuki–Miyaura cross-coupling reaction of 9-bromoanthracene with pyridin-4-ylboronic acid and purified by column chromatography on silica gel. Light-yellow crystals of 4-(anthracen-9-yl)-pyridine suitable for X-ray diffraction were collected by the solvent evaporation method. In the crystal, pairs of molecules are connected by intermolecular C—H...π (pyridine) interactions [d(H7...Cg) = 2.7391 (2) Å], forming cyclic centrosymmetric dimers, further resulting in an infinite one-dimensional linear chain along the c-axis direction. Weak face-to-face π–π stacking interactions [d(Cg...Cg) = 3.6061 (2) Å] link neighboring lamellar networks into the supramolecular structure.

Recently, immune response modulation at the epigenetic level is illustrated in studies, but the possible function of RNA 5-methylcytosine (m5C) modification in cell infiltration within the tumor microenvironment (TME) is still unclear. Three different m5C modification patterns were identified, and high differentiation degree was observed in the cell infiltration features within TME under the above three identified patterns. A low m5C-score, which was reflected in the activated immunity, predicted the relatively favorable prognostic outcome. A small amount of effective immune infiltration was seen in the high m5C-score subtype, indicating the dismal patient survival. Our study constructed a diagnostic model using the 10 signature genes highly related to the m5C-score, discovered that the model exhibited high diagnostic accuracy for PTC, and screened out five potential drugs for PTC based on this m5C-score model. m5C modification exerts an important part in forming the TME complexity and diversity. It is valuable to evaluate the m5C modification patterns in single tumors, so as to enhance our understanding towards the infiltration characterization in TME.

Abstract Background Corynebacterium glutamicum is a traditional food-grade industrial microorganism, in which an efficient endotoxin-free recombinant protein expression factory is under developing in recent years. However, the intrinsic disadvantage of low recombinant protein expression level is still difficult to be solved. Here, according to the bacteria-specific polycistronic feature that multiple proteins can be translated in one mRNA, efforts have been made to insert a leading peptide gene upstream of target genes as an expression enhancer, and it is found that this can remarkably improve the expression level of proteins under the control of inducible tac promoter in C. glutamicum. Results In this research, the Escherichia coli (E. coli) tac promoter combined with 24 different fore-cistron sequences were constructed in a bicistronic manner in C. glutamicum. Three strong bicistronic expression vectors were isolated and exhibited high efficiency under different culture conditions. The compatibility of these bicistronic vectors was further validated using six model proteins- aldehyde dehydrogenase (ALDH), alcohol dehydrogenase (ADH), RamA (regulator of acetate metabolism), Bovine interferon-α (BoIFN-α), glycoprotein D protein (gD) of infectious bovine rhinotracheitis virus (IBRV) and procollagen type Ι N-terminal peptide (PΙNP). All examined proteins were highly expressed compared with the original vector with tac promoter. Large-scale production of PΙNP was also performed in fed-batch cultivation, and the highest PΙNP production level was 1.2 g/L. Conclusion In this study, the strength of the inducible tac promoter for C. glutamicum was improved by screening and inserting fore-cistron sequences in front of the target genes. Those vectors with bicistronic expression patterns have strong compatibility for expressing various heterogeneous proteins in high yield. This new strategy could be used to further improve the performance of inducible promoters, achieving double competence of inducible control and high yield.

Abstract While biochar soil amendment has been widely proposed as a soil organic carbon (SOC) sequestration strategy to mitigate detrimental climate changes in global agriculture, the SOC sequestration was still not clearly understood for the different effects of fresh and aged biochar on SOC mineralization. In the present study of a two‐factorial experiment, topsoil samples from a rice paddy were laboratory‐incubated with and without fresh or aged biochar pyrolyzed of wheat residue and with and without crop residue‐derived dissolved organic matter (CRM) for monitoring soil organic matter decomposition under controlled conditions. The six treatments included soil with no biochar, with fresh biochar and with aged biochar treated with CRM, respectively. For fresh biochar treatment, the topsoil of a same rice paddy was amended with wheat biochar directly from a pyrolysis wheat straw, the soil with aged biochar was collected from the same soil 6 years following a single amendment of same biochar. Total CO2 emission from the soil was monitored over a 64 day time span of laboratory incubation, while microbial biomass carbon and phospholipid fatty acid (PLFA) were determined at the end of incubation period. Without CRM, total organic carbon mineralization was significantly decreased by 38.8% with aged biochar but increased by 28.9% with fresh biochar, compared to no biochar. With CRM, however, the significantly highest net carbon mineralization occurred in the soil without biochar compared to the biochar‐amended soil. Compared to aged biochar, fresh biochar addition significantly increased the total PLFA concentration by 20.3%–33.8% and altered the microbial community structure by increasing 17:1ω8c (Gram‐negative bacteria) and i17:0 (Gram‐positive bacteria) mole percentages and by decreasing the ratio of fungi/bacteria. Furthermore, biochar amendment significantly lowered the metabolic quotient of SOC decomposition, thereby becoming greater with aged biochar than with fresh biochar. The finding here suggests that biochar amendment could improve carbon utilization efficiency by soil microbial community and SOC sequestration potential in paddy soil can be enhanced by the presence of biochar in soil over the long run.

Abstract Background In terms of protein production, the internal environment of the host influences the activity of expression elements, thus affecting the expression level of the target protein. Native expression elements from a specific strain always function well in the original host. In the present study, to enhance the endoxylanase (XynA) production level in Corynebacterium glutamicum CGMCC1.15647 with its native expression elements, approaches to reduce host expression obstacles and to promote expression were evaluated. Results We identified the signal peptide of CspB2 in C. glutamicum CGMCC1.15647 by MALDI-TOF and applied it along with its promoter for the production of endoxylanase (XynA) in this strain. The native cspB2 promoter and cspB2 signal peptide are superior to the well-used cspB1 promoter and cspA signal peptide for XynA expression in C. glutamicum CGMCC1.15647, and expression in this strain is superior to the expression in C. glutamicum ATCC13032. The highest XynA secretion efficiency level in deep 24-well plates level (2492.88 U/mL) was achieved by disruption of the cell wall protein CspB2 and the protease ClpS, chromosomal integration of xynA and coexisting plasmid expression, which increased expression 11.43- and 1.35-fold compared to that of chromosomal expression and pXMJ19-xynA-mediated expression in the original strain, respectively. In fed-batch cultivation, the highest XynA accumulation (1.77 g/L) was achieved in the culture supernatant after 44 h of cultivation. Conclusion Adaptation between the expression elements and the host is crucial for XynA production in C. glutamicum CGMCC1.15647. Strategies including host optimization, chromosomal integration, and coexistence of plasmids were useful for efficient protein production in C. glutamicum.

A chemiluminescent immunoassay for human serum osteocalcin, or bone Gla protein, was established using a double-antibody sandwich model. Examination of the hook effect revealed that it was significantly reduced, and no hook effect was observed at an osteocalcin concentration of 4000 ng/mL. The established method showed good analytical performance and thermal stability. The limit of detection was 0.03 ng/mL. The intra-assay coefficient of variation (CV) was 3.22%–5.64%, the interassay CV was 4.42%–7.25%, and the recovery rate was 93.22%–107.99%. Cross-reactivity (CR) was not observed with bovine, rat, mouse, rabbit, or porcine samples. The developed method showed a good correlation with the N-MID product from Roche. In total, 1069 clinical patient samples were measured using the reagent kit developed in this study.

Plant defense: Regulating resistance to ring rot A gene with multiple functions in defense against apple ring rot may provide a target for breeding apple varieties resistant to this devastating and enigmatic fungal disease. In China, the world’s greatest producer of apples, apple ring rot poses a severe challenge to growers. A Chinese team led by Shujing Wu and Xuesen Chen, of Shandong Agricultural University, explored the role of a gene, MdSYP121, thought to provide resistance to ring rot. A combination of genetic, genomic, and transgenic experiments showed that MdSYP121 does indeed confer resistance to the disease, and may do this both by regulating hormonal pathways of defense and by stimulating enzymes involved in defensive oxidation/reduction reactions. Overall, their results suggest potential foci for breeding programmes aimed at reducing the susceptibility of apple cultivars to the ring rot fungus.

It is widely accepted that features such as pI, length, molecular mass and amino acid (AA) sequence have a significant influence on protein solubility. Here, we mainly focused on AA composition and explored those that most affected the soluble expression level of human serum albumin (HSA) domain antibody (dAb). The soluble expression and sequence of 65 dAb variants were analysed using clustering and linear modelling. Certain AAs significantly affected the soluble expression level of dAb, with the specific AA combinations being (S, R, N, D, Q), (G, R, C, N, S) and (R, S, G); these combinations respectively affected the dAb expression level in the broth supernatant, the level in the pellet lysate and total soluble dAb. Among the 20 AAs, R displayed a negative influence on the soluble expression level, whereas G and S showed positive effects. A linear model was built to predict the soluble expression level from the sequence; this model had a prediction accuracy of 80 %. In summary, increasing the content of polar AAs, especially G and S, and decreasing the content of R, was helpful to improve the soluble expression level of HSA dAb.

Abstract Background Corynebacterium glutamicum (C. glutamicum) has traditionally been used as a microbial cell factory for the industrial production of many amino acids and other industrially important commodities. C. glutamicum has recently been established as a host for recombinant protein expression; however, some intrinsic disadvantages could be improved by genetic modification. Gene editing techniques, such as deletion, insertion, or replacement, are important tools for modifying chromosomes. Results In this research, we report a CRISPR/Cas9 system in C. glutamicum for rapid and efficient genome editing, including gene deletion and insertion. The system consists of two plasmids: one containing a target-specific guide RNA and a homologous sequence to a target gene, the other expressing Cas9 protein. With high efficiency (up to 100%), this system was used to disrupt the porB, mepA, clpX and Ncgl0911 genes, which affect the ability to express proteins. The porB- and mepA-deletion strains had enhanced expression of green fluorescent protein, compared with the wild-type stain. This system can also be used to engineer point mutations and gene insertions. Conclusions In this study, we adapted the CRISPR/Cas9 system from S. pyogens to gene deletion, point mutations and insertion in C. glutamicum. Compared with published genome modification methods, methods based on the CRISPR/Cas9 system can rapidly and efficiently achieve genome editing. Our research provides a powerful tool for facilitating the study of gene function, metabolic pathways, and enhanced productivity in C. glutamicum.

Corynebacterium glutamicum (C. glutamicum) is a favorable host cell for the production of recombinant proteins, such as important enzymes and pharmaceutical proteins, due to its excellent potential advantages. Herein, we sought to systematically explore the influence of recombinant protein expression on the transcription and metabolism of C. glutamicum. Two C. glutamicum strains, the wild-type strain and an engineered strain expressing enhanced green fluorescent protein (EGFP), were cultured in parallel in 5-L bioreactors to study the change in metabolism in the process of EGFP expression. The results revealed that EGFP expression had great effects on the growth and metabolism of C. glutamicum and contributed to metabolism-like anaerobic conditions as follows: glycolysis was enhanced, the TCA cycle was shunted, and Glu, Val, Met, lactate and acetate were accumulated to produce sufficient ATP for EGFP production and transfer. Many differentially expressed genes related to ribosomal protein, transcriptional regulators, and energy metabolism were found to be expressed in the presence of EGFP, laying the foundation for identifying genomic loci to change the flow of the host cell metabolism to improve the ability of expressing foreign proteins in C. glutamicum.

Dissolved oxygen (DO) is an important factor in the fermentation process of Corynebacterium glutamicum, which is a widely used aerobic microbe in bio-industry. Herein, we described RNA-seq for C. glutamicum under different DO levels (50%, 30% and 0%) in 5 L bioreactors. Multivariate data analysis (MVDA) models were used to analyze the RNA-seq and metabolism data to investigate the global effect of DO on the transcriptional distinction of the substance and energy metabolism of C. glutamicum. The results showed that there were 39 and 236 differentially expressed genes (DEGs) under the 50% and 0% DO conditions, respectively, compared to the 30% DO condition. Key genes and pathways affected by DO were analyzed, and the result of the MVDA and RNA-seq revealed that different DO levels in the fermenter had large effects on the substance and energy metabolism and cellular redox balance of C. glutamicum. At low DO, the glycolysis pathway was up-regulated, and TCA was shunted by the up-regulation of the glyoxylate pathway and over-production of amino acids, including valine, cysteine and arginine. Due to the lack of electron-acceptor oxygen, 7 genes related to the electron transfer chain were changed, causing changes in the intracellular ATP content at 0% and 30% DO. The metabolic flux was changed to rebalance the cellular redox. This study applied deep sequencing to identify a wealth of genes and pathways that changed under different DO conditions and provided an overall comprehensive view of the metabolism of C. glutamicum. The results provide potential ways to improve the oxygen tolerance of C. glutamicum and to modify the metabolic flux for amino acid production and heterologous protein expression.

WDR5 is a core component of the human mixed lineage leukemia-2 complex, which plays central roles in ER positive tumour cells and is a major driver of androgen-dependent prostate cancer cell proliferation. Given the similarities between breast and prostate cancers, we explore the potential prognostic value of WDR5 gene expression on breast cancer survival. Our findings reveal that WDR5 over-expression is associated with poor breast cancer clinical outcome in three gene expression data sets and BreastMark. The eQTL analysis reveals 130 trans-eQTL SNPs whose genes mapped with statistical significance are significantly associated with patient survival. These genes together with WDR5 are enriched with "cellular development, gene expression, cell cycle" signallings. Knocking down WDR5 in MCF7 dramatically decreases cell viability, but does not alter tumour cell response to doxorubicin. Our study reveals the prognostic value of WDR5 expression in breast cancer which is under long-range regulation of genes involved in cell cycle, and anthracycline could be coupled with treatments targeting WDR5 once such a regimen is available.