Array Point Discharge as Enhanced Tandem Excitation Source for Miniaturized Optical Emission Spectrometer.

A new compact tandem excitation source was designed and constructed by using an array point discharge (ArrPD) microplasma for a miniaturized optical emission spectrometer through coupling a hydride generation (HG) unit as a sample introduction device. Three pairs of point discharges were arranged in sequence in a narrow discharge chamber to construct the ArrPD microplasma, for improved excitation capability owing to the serial excitation. Besides, the discharge plasma region was greatly enlarged, therefore, more gaseous analytes could be intercepted to enter into the microplasma for sufficient excitation, for improved excitation efficiency and OES signal. To better understand the effectiveness of the proposed ArrPD source, a new instrument for simultaneous detection of atomic emission and absorption spectral responses was also proposed, designed, and constructed to reveal the excitation and enhancement process in the discharge chamber. Under the optimized conditions, the limits of detection (LODs) of As, Ge, Hg, Pb, Sb, Se, and Sn were 0.7, 0.4, 0.05, 0.7, 0.3, 2, and 0.08 µg L-1, respectively, and the relative standard deviations (RSDs) were all less than 4%. Compared with a commonly used single point discharge microplasma source, the analytical sensitivities of these seven elements were improved by 3-6-fold. Certified Reference Materials (CRMs) were successfully analyzed with this miniaturized spectrometer, which features low power, compactness, portability, and high detectability, and is thereby a great prospect in the field of elemental analytical chemistry.

Residual periodontal ligament in the extraction socket promotes the dentin regeneration potential of DPSCs in the rabbit jaw.

BACKGROUND: Because of the low regeneration efficiency and unclear underlying molecular mechanism, tooth regeneration applications are limited. In this study, we explored the influence of residual periodontal ligament on the dentin regeneration potential of dental pulp stem cells (DPSCs) in the jaw. METHODS: To establish a tooth regeneration model, the incisors of New Zealand white rabbits were extracted while preserving residual periodontal ligament, followed by the implantation of DPSCs. After 3 months, micro-computed tomography (micro-CT), stereomicroscopy and scanning electron microscopy (SEM) were used to observe the volume, morphology and microstructure of regenerated tissue. Histological staining and immunostaining analyses were used to observe the morphological characteristics and expression of the dentin-specific proteins DMP1 and DSPP. To explore the mechanism, DPSCs and periodontal ligament stem cells (PDLSCs) were cocultured in vitro, and RNA was collected from the DPSCs for RNA-seq and bioinformatic analysis. RESULTS: The results of micro-CT and stereomicroscopy showed that the number of sites with regeneration and the volume of regenerated tissue in the DPSCs/PDL group (6/8, 1.07 ± 0.93 cm3) were larger than those in the DPSCs group (3/8, 0.23 ± 0.41 cm3). The results of SEM showed that the regenerated dentin-like tissue in the DPSCs and DPSCs/PDL groups contained dentin tubules. Haematoxylin and eosin staining and immunohistochemical staining indicated that compared with the DPSCs group, the DPSCs/PDL group showed more regular regenerated tissue and higher expression levels of the dentin-specific proteins DMP1 and DSPP (DMP1: P = 0.02, DSPP: P = 0.01). RNA-seq showed that the coculture of DPSCs with PDLSCs resulted in the DPSCs differentially expressing 427 mRNAs (285 upregulated and 142 downregulated), 41 lncRNAs (26 upregulated and 15 downregulated), 411 circRNAs (224 upregulated and 187 downregulated), and 19 miRNAs (13 upregulated and 5 downregulated). Bioinformatic analysis revealed related Gene Ontology function and signalling pathways, including extracellular matrix (ECM), tumour necrosis factor (TNF) signalling and chemokine signalling pathways. CONCLUSIONS: Residual periodontal ligament in the extraction socket promotes the dentin regeneration potential of DPSCs in the jaw. RNA-seq and bioinformatic analysis revealed that ECM, TNF signalling and chemokine signalling pathways may represent the key factors and signalling pathways.

Bibliometric and visualized analysis of scientific publications on rehabilitation of rotator cuff injury based on web of science.

Background: Since the discovery of rehabilitation as an intervention for rotator cuff injury, its impact on the recovery of rotator cuff injury has attracted crucial attention, and the number of related studies is increasing worldwide. There were no bibliometric and visualized analysis studies in this field. This study aimed to investigate the research hotpots and trends in the rehabilitation of rotator cuff injury via bibliometric and visualized analysis and to identify the future development of clinical practice. Method: The publications regarding rehabilitation of rotator cuff injury from inception to December 2021 were obtained from the Web of Science Core Collection database. The trends of publications, co-authorship and co-occurrence analysis and visualized analysis were carried out using Citespace, VOSviewer, Scimago Graphica software, and R Project. Results: A total of 795 publications were included in this study. The number of publications significantly increased yearly. The United States published the highest number of related papers and the papers published by the United States had the highest citations. The University of Laval, the University of Montreal and Keele University were the top 3 most contributive institutions. Additionally, the Journal of Shoulder and Elbow Surgery was the journal with the highest number of publications. The most common keywords were "rotator cuff", "rehabilitation", "physical therapy", "management", and "telerehabilitation". Conclusion: The total number of publications has shown a steady upward trend. The cooperation between countries globally was still relatively lacking, and therefore it is necessary to strengthen cooperation between different countries and regions to provide conditions for multi-center, large sample, and high-quality research. In addition to the relatively mature rehabilitation of rotator cuff injury such as passive motion or exercise therapy, telerehabilitation has also attracted much attention with the progress of science.

Molecular Collapse States in Graphene/WSe_{2} Heterostructure Quantum Dots.

In relativistic physics, both atomic collapse in a heavy nucleus and Hawking radiation in a black hole are predicted to occur through the Klein tunneling process that couples particles and antiparticles. Recently, atomic collapse states (ACSs) were explicitly realized in graphene because of its relativistic Dirac excitation with a large "fine structure constant." However, the essential role of the Klein tunneling in the ACSs remains elusive in experiment. Here we systematically study the quasibound states in elliptical graphene quantum dots (GQDs) and two coupled circular GQDs. Bonding and antibonding molecular collapse states formed by two coupled ACSs are observed in both systems. Our experiments supported by theoretical calculations indicate that the antibonding state of the ACSs will change into a Klein-tunneling-induced quasibound state revealing deep connection between the ACSs and the Klein tunneling.

Intestinal prolapse and exposure after peritoneal dialysis in low-birth-weight preterm infants with acute renal failure: a case report.

Background: The mortality rate of acute kidney injury (AKI) in low-birth-weight premature infants with acute renal failure is extremely high. Since small hemodialysis catheters do not exist, peritoneal dialysis (PD) is the most suitable dialysis method. At present, only a few studies have reported cases of PD in low-birth-weight newborns. Case Description: On September 8, 2021, a 10-day-old, low-birth-weight preterm infant, who presented with neonatal respiratory distress syndrome and acute renal failure, was admitted to the Second Affiliated Hospital of Kunming Medical University, China. The patient was the elder of twins and had experienced acute renal failure, hyperkalemia, and anuria following the onset of respiratory distress syndrome. During the initial PD catheterization operation, a double Tenckhoff adult PD catheter cut short by 2 cm was used, with the inner cuff placed in the skin. However, the surgical incision was relatively large, and PD fluid leakage occurred. Later, the incision tore, and the intestines prolapsed when the patient cried. The intestines were returned to the abdominal cavity in an emergency operation, and the PD catheter was placed again. This time, the inner Tenckhoff cuff was placed outside the skin, and PD fluid leakage did not reoccur. However, the patient also experienced a decrease in heart rate and blood pressure, as well as severe pneumonia and peritonitis. Following an active rescue, the patient recovered well. Conclusions: The PD method effectively treats low-birth-weight preterm neonates with AKI. An adult Tenckhoff catheter was shortened by 2 cm and successfully used in the PD treatment of a low-birth-weight preterm infant. However, the catheter placement should be outside the skin, and the incision should be as small as possible to avoid leakage and incision tears.

Evolutionary Analysis of Respiratory Burst Oxidase Homolog (RBOH) Genes in Plants and Characterization of ZmRBOHs.

The respiratory burst oxidase homolog (RBOH), as the key producer of reactive oxygen species (ROS), plays an essential role in plant development. In this study, a bioinformatic analysis was performed on 22 plant species, and 181 RBOH homologues were identified. A typical RBOH family was identified only in terrestrial plants, and the number of RBOHs increased from non-angiosperms to angiosperms. Whole genome duplication (WGD)/segmental duplication played a key role in RBOH gene family expansion. Amino acid numbers of 181 RBOHs ranged from 98 to 1461, and the encoded proteins had molecular weights from 11.1 to 163.6 kDa, respectively. All plant RBOHs contained a conserved NADPH_Ox domain, while some of them lacked the FAD_binding_8 domain. Plant RBOHs were classified into five main subgroups by phylogenetic analysis. Most RBOH members in the same subgroup showed conservation in both motif distribution and gene structure composition. Fifteen ZmRBOHs were identified in maize genome and were positioned in eight maize chromosomes. A total of three pairs of orthologous genes were found in maize, including ZmRBOH6/ZmRBOH8, ZmRBOH4/ZmRBOH10 and ZmRBOH15/ZmRBOH2. A Ka/Ks calculation confirmed that purifying selection was the main driving force in their evolution. ZmRBOHs had typical conserved domains and similar protein structures. cis-element analyses together with the expression profiles of the ZmRBOH genes in various tissues and stages of development suggested that ZmRBOH was involved in distinct biological processes and stress responses. Based on the RNA-Seq data and qRT-PCR analysis, the transcriptional response of ZmRBOH genes was examined under various abiotic stresses, and most of ZmRBOH genes were up-regulated by cold stress. These findings provide valuable information for further revealing the biological roles of ZmRBOH genes in plant development and abiotic stress responses.

Transmembrane Protein 68 Functions as an MGAT and DGAT Enzyme for Triacylglycerol Biosynthesis.

Triacylglycerol (TG) biosynthesis is an important metabolic process for intracellular storage of surplus energy, intestinal dietary fat absorption, attenuation of lipotoxicity, lipid transportation, lactation and signal transduction in mammals. Transmembrane protein 68 (TMEM68) is an endoplasmic reticulum (ER)-anchored acyltransferase family member of unknown function. In the current study we show that overexpression of TMEM68 promotes TG accumulation and lipid droplet (LD) formation in a conserved active sites-dependent manner. Quantitative targeted lipidomic analysis showed that diacylglycerol (DG), free fatty acid (FFA) and TG levels were increased by TMEM68 expression. In addition, TMEM68 overexpression affected the levels of several glycerophospholipids, such as phosphatidylcholine, phosphatidylethanolamine and phosphatidylinositol, as well as sterol ester contents. TMEM68 exhibited monoacylglycerol acyltransferase (MGAT) and diacylglycerol acyltransferase (DGAT) activities dependent on the conserved active sites in an in vitro assay. The expression of lipogenesis genes, including DGATs, fatty acid synthesis-related genes and peroxisome proliferator-activated receptor γ was upregulated in TMEM68-overexpressing cells. These results together demonstrate for the first time that TMEM68 functions as an acyltransferase and affects lipogenic gene expression, glycerolipid metabolism and TG storage in mammalian cells.

Discovery and optimization of olanzapine derivatives as new ferroptosis inhibitors.

Ferroptosis is a new type of cell death associated with many human diseases. It is a new strategy to discover ferroptosis inhibitors for the treatment of ferroptosis-related diseases. Here the FDA-approved drug library containing 1160 molecules was screened for ferroptosis inhibitors in RSL3-induced HT22 mouse hippocampal neuronal cells. As a result, olanzapine showed potent ferroptosis inhibitory activity (EC50 = 1.18 µM). Structural optimization and the structure-activity relationships (SARs) analysis led to the synthesis of 41 new derivatives (4-44) and one known compound 45. Comparing with olanzapine, its derivative 36 showed nearly sixteen-folds improved ferroptosis inhibition and low cytotoxicity (EC50 = 0.074 µM, CC50 = 18.8 µM). Further mechanistic studies revealed that compound 36 specifically inhibited ferroptosis by its antioxidative ability. This work demonstrates that olanzapine protected RSL3-induced ferroptosis in HT22 cell, and its derivative 36 having nanomolar ferroptosis inhibitory activity merit to be developed for drugs against ferroptosis-related neurological diseases.

Chemical constituents of Ajuga forrestii and their anti-ferroptosis activity.

Six new neoclerodane diterpenoids (1-6), along with ten known compounds (7-16), were isolated from Ajuga forrestii. Their structures were elucidated by HRESIMS, 1D and 2D NMR, ECD calculation, and single-crystal X-ray diffraction analysis. The structure of a known neoclerodane diterpene ajudecunoid C (6) was revised based on the reported NMR empirical rules. All the isolates were evaluated for their inhibitory effect on RSL3-induced ferroptosis in HT22 mouse hippocampal neuronal cells. Among them, compounds 8, 9, and 12 significantly inhibited RSL3-induced ferroptosis with EC50 values of 0.45 µM, 0.076 µM, and 0.14 µM.

Clinical characterization of Lamb-Shaffer syndrome: a case report and literature review.

BACKGROUND: Lamb-Shaffer syndrome (LAMSHF, MIM 616,803) is a rare neurodevelopmental disorder due to haploinsufficiency of SOX5. Furthermore, studies about the clinical features of LAMSHF patients with same allele of c.1477C > T (p. R493*) are very limited. CASE PRESENTATION: We analyzed the phenotypes of one of our cases and two previously reported cases with c.1477C > T (p. R493*), and reviewed the correlating literature. A de novo heterozygous variation c.1477C > T (p. R493*) in SOX5 was identified in a 4 years and 2 months old boy with global development delay by trio-based whole exome sequencing. We compared our case and previously 2 cases reported with recurrent variation, the overlapping clinical features are global developmental delay or intellectual disability, language delay and scoliosis, but their other clinical characteristics are different. CONCLUSIONS: This study suggests that the clinical features of LAMSHF patients with recurrent variations in the SOX5 gene are different. It is suggested that the LAMSHF-related SOX5 gene should be screened and included as one of the candidate genes for neurodevelopmental disorders of unknown etiology.

The association between liver fibrosis and depression in patients after ischemic stroke.

BACKGROUND AND OBJECTIVE: Liver fibrosis has been considered a predictor of cardiovascular disease. This study aimed to evaluate whether the degree of liver fibrosis is related to post-stroke depression (PSD) at 3 months follow-up. METHODS: We prospectively and continuously enrolled patients with first-ever ischemic stroke from June 2020 to January 2022. Liver fibrosis was measured after admission by calculating the Fibrosis-4 index (FIB-4) and stratified into two categories (< 2.67 versus ≥ 2.67). Patients with a 17-item Hamilton Depression Scale score > 7 were further evaluated using the Chinese version of the structured clinical interview of DSM-IV, for diagnosing PSD at 3 months. RESULTS: A total of 326 patients (mean age 66.6 years, 51.5% male) were recruited for the study. As determined by the FIB-4 score, 80 (24.5%) patients had advanced liver fibrosis. During the follow-up, PSD was observed in 91 patients, which accounted for 27.9% (95% confidence interval [CI] 25.5%-30.5%) of the cohort. The prevalence of advanced liver fibrosis was higher in PSD patients than those without PSD (40.0% versus 24.0%; P = 0.006). After adjustment for covariates in the multivariate logistic analysis, advanced fibrosis was significantly associated with PSD (odds ratio [OR], 1.88; 95% CI, 1.03-3.42; P = 0.040). Similar results were found when the FIB-4 was analyzed as a continuous variable. CONCLUSIONS: This study found that advanced liver fibrosis was associated with an increased risk of 3-month PSD. FIB-4 score may be valuable for screening depressive symptoms in ischemic stroke patients.

Development of the Droplet Digital PCR Method for the Detection and Quantification of Erwinia pyrifoliae.

Black shoot blight disease caused by Erwinia pyrifoliae has serious impacts on quality and yield in pear production in Korea; therefore, rapid and accurate methods for its detection are needed. However, traditional detection methods require a great deal of time and fail to achieve absolute quantification. In the present study, we developed a droplet digital polymerase chain reaction (ddPCR) method for the detection and absolute quantification of E. pyrifoliae using a pair of species-specific primers. The detection range was 103 - 107 copies/ml (DNA templates) and cfu/ml (cell culture templates). This new method exhibited good linearity and repeatability and was validated by absolute quantification of E. pyrifoliae DNA copies from samples of artificially inoculated immature pear fruits. Here, we present the first study of ddPCR assay for the detection and quantification of E. pyrifoliae. This method has potential applications in epidemiology and for the early prediction of black shoot blight outbreaks.

The mechanism of coumarin inhibits germination of ryegrass (Lolium perenne) and its application as coumarin-carbon dots nanocomposites.

BACKGROUND: As an important plant allelochemical, coumarin can effectively inhibit the germination of various seeds. However, little is known about the inhibition mechanism of coumarin on weed seed germination. Moreover, the herbicidal activity of coumarin is needed to be improved as a natural pesticide. RESULTS: Coumarin had the highest inhibition effect on the ryegrass (Lolium perenne) seed, where coumarin disturbed the hormone pathway by decreasing the content of gibberellic acid 3, resulting in the reduction of amylase activity and consumption of starch during the germination process of ryegrass seed. Moreover, coumarin induced decreased activity of catalase and subsequently led to the accumulation of hydrogen peroxide and malondialdehyde, causing oxidative stress during the germination process of ryegrass seed. Furthermore, to enhance the herbicidal activity of coumarin, carbon dots (CDs) modified with polyetherimide were prepared, characterized, and then combined with coumarin to form coumarin-carbon dots (Cm-CDs) nanocomposites. Compared with coumarin, Cm-CDs nanocomposites significantly increased the herbicidal activity of coumarin on ryegrass, which implies that Cm-CDs nanocomposites could be used as a potential formulation to improve the herbicidal activity of coumarin. CONCLUSION: This study not only reveals the mechanism of coumarin on ryegrass germination, but also develop Cm-CDs nanocomposites to enhance the herbicidal activity of coumarin. Our findings will stimulate the application of Cm-CDs nanomaterials as an effective and environmentally friendly formulation in agriculture. © 2023 Society of Chemical Industry.

Consumer Preference for Virtual Reality Advertisements with Human-Scene Interaction: An Intermediary Based on Psychological Needs.

Consumption is undergoing a digital revolution brought about by virtual technology, and the emergence of virtual spokespersons has changed the pattern of advertising endorsement sales. Are virtual character spokespersons better than real celebrity spokespersons? This article aims to explore the relationships between different types of advertising spokespersons, different types of advertising scenarios, and consumers' purchase intentions. The results of 2 pilot experiments and 4 formal experiments show that compared with spokespersons by real people, virtual character spokespersons have a greater positive impact on consumers' purchase intentions, which is achieved through the mediating role of psychological need satisfaction. In addition, different advertising scenarios provided by virtual technology have a moderating effect on this process. This empirical study investigates the possible positive effects of virtual character spokesperson compared with real character spokesperson and expands the results of spokesperson category based on self-determination theory.

Real-Space Mapping of Local Subdegree Lattice Rotations in Low-Angle Twisted Bilayer Graphene.

In two-dimensional small-angle twisted bilayers, van der Waals (vdW) interlayer interaction introduces an atomic-scale reconstruction, which consists of a moiré-periodic network of local subdegree lattice rotations. However, real-space measurement of the subdegree lattice rotation requires extremely high spatial resolution, which is an outstanding challenge in an experiment. Here, a topmost small-period graphene moiré pattern is introduced as a magnifying lens to magnify sub-Angstrom lattice distortions in small-angle twisted bilayer graphene (TBG) by about 2 orders of magnitude. Local moiré periods of the topmost graphene moiré patterns and low-energy van Hove singularities of the system are spatially modified by the atomic-scale reconstruction of the underlying TBG, thus enabling real-space mapping of the networks of the subdegree lattice rotations both in structure and in electronic properties. Our results indicate that it is quite facile to study subdegree lattice rotation in vdW systems by measuring the periods of the topmost moiré superlattice.

Molecular Basis for the Selectivity of the Succinate Dehydrogenase Inhibitor Cyflumetofen between Pest and Predatory Mites.

Acaricides that act as inhibitors of the mitochondrial succinate dehydrogenase (SDHIs) provide excellent control of phytophagous mites but display limited toxicity to predatory mites and other beneficial organisms. However, the molecular mechanism of selectivity is not fully understood. Here, we first confirm that SDHI acaricides are over 10,000-fold more toxic to spider mites than predatory mites. Next, we show that differential penetration, pro-acaricide activation, or metabolism are most likely not the main reason for this selectivity. In contrast, the inhibition of AB-1 on the SDH target is approximately 200-fold more potent in spider mites compared to predatory mites, revealing strong target-site selectivity. Strikingly, a key motif associated with differential binding was identified and validated by gene editing in Drosophila. Our findings contribute to understanding the selectivity of SDHIs, which can be used for the rational design of selective acaricides in support of an integrated pest management.

Leonurine promotes the maturation of healthy donors and multiple myeloma patients derived-dendritic cells via the regulation on arachidonic acid metabolism.

Objective: Leonurine is a bioactive alkaloid compound extracted from Leonurus japonicus Houtt, which potentially has immunomodulatory effects. The immunomodulatory effect and mechanism of leonurine on monocyte derived dendritic cells (moDCs) from healthy donors (HDs) and multiple myeloma (MM) patients were investigated for the first time. Methods: Peripheral blood from HDs and MM patients was isolated for peripheral blood mononuclear cells (PBMCs). The generation of moDCs was conducted by the incubation of monocytes from PBMCs in the medium consisting of RPMI 1640 medium, 2 mmol/L L-glutamine, 5% human serum, 800 U/mL GM-CSF, 500 U/mL IL-4, 100 U/mL penicillin and 0.1 mg/mL streptomycin. During the incubation of 7 days, the cells were administrated with 1 µM leonurine or 1 × PBS as the control group. On the 8th day, cells were harvested. The expression of maturation associated surface markers CD40, CD83, and HLA-DR on moDCs was analyzed by flow cytometry. Moreover, moDCs with or without 1 µM leonurine administration were evaluated by LC-MS/MS for metabolomics which was further analyzed for the potential mechanism of leonurine on moDCs. Results: The proportion of moDCs in the harvested cells was significantly higher in the HD group (n = 14) than in the MM patient group (n = 11) (p = 0.000). Leonurine significantly enhanced the median fluorescence intensity of CD83, HLA-DR and CD40 expression on HD-moDCs (n = 14; p = 0.042, p = 0.013, p = 0.084) as well as MM paitent-moDCs (n = 11; p = 0.020, p = 0.006, p = 0.025). The metabolomics data showed that in moDCs (HD, n = 15), 18 metabolites in the pathway of arachidonic acid metabolism showed significant differences between the leonurine group and the control group (VIP all >1 and P all <0.05). To be specific, 6-Keto-PGE1, 8,9-DHET, 11 (R)-HETE, 12-Keto-LTB4, 12-OxoETE, 15 (S)-HETE, 15-Deoxy-Delta12,14-PGJ2, 15-Keto-PGF2a, 20-COOH-LTB4, Lecithin, PGA2, PGB2, PGE2, PGF2a, PGG2, Prostacyclin were significantly upregulated in the leonurine group than in the control group, while Arachidonic Acid and TXB2 were significantly downregulated in the leonurine group than in the control group. Conclusion: Leonurine significantly promotes the maturation of moDCs derived from HDs and MM patients, the mechanism of which is related to arachidonic acid metabolism.

Life-threatening pulmonary coinfection with Mycobacterium tuberculosis and Aspergillus lentulus in a diabetic patient diagnosed by metagenome next-generation sequencing.

BACKGROUND: China has a double burden of diabetes mellitus and tuberculosis. Diabetes mellitus and tuberculosis are both important risk factors for Aspergillus infection. Aspergillus lentulus is an emerging fungal pathogen in China and invasive aspergillosis due to A. lentulus is associated with high mortality. CASE PRESENTATION: A 79-year-old man was admitted to our hospital, complaining of a 7-day history of fever. Five days before admission, he was diagnosed with pulmonary infection at a local hospital, but his symptoms did not relieve after antibiotic therapy. The patient was diagnosed with diabetes mellitus two months ago. About 20 days ago, he began to present chest tightness and shortness of breath after physical activity. After admission, he developed continuous fever and rapid respiratory deterioration, and finally died after his family abandoned treatment. Pulmonary coinfection with M. tuberculosis and A. lentulus was identified by metagenome next-generation sequencing (mNGS) from bronchoalveolar lavage fluid. CONCLUSIONS: Clinicians and laboratories should be alert to the emerging A. lentulus infection in China due to its drug-resistance and high mortality. In comparison with conventional methods, mNGS has a great advantage for the diagnosis of mixed pulmonary infection.

Using the inner membrane of Escherichia coli as a scaffold to anchor enzymes for metabolic flux enhancement.

Clustering enzymes in the same metabolic pathway is a natural strategy to enhance productivity. Synthetic protein, RNA and DNA scaffolds have been designed to artificially cluster multiple enzymes in the cell, which require complex construction processes and possess limited slots for target enzymes. We utilized the Escherichia coli inner cell membrane as a native scaffold to cluster four fatty acid synthases (FAS) and achieved to improve the efficiency of fatty acid synthesis in vivo. The construction strategy is as simple as fusing target enzymes to the N-terminus or C-terminus of the membrane anchor protein (Lgt), and the number of anchored enzymes is not restricted. This novel device not only presents a similar efficiency in clustering multiple enzymes to that of other artificial scaffolds but also promotes the product secretion, driving the entire metabolic flux forward and further increasing the gross yield compared with that in a cytoplasmic scaffold system.