Aims
We aimed to explore Yarrowia lipolytica carbonyl reductases as effective biocatalysts and to develop efficient asymmetric reduction systems for chiral alcohol synthesis.
Methods and Results
...Yarrowia lipolytica carbonyl reductase genes were obtained via homologous sequence amplification strategy. Two carbonyl reductases, YaCRI and YaCRII, were identified and characterized, and used to catalyse the conversion of 2‐hydroxyacetophenone (2‐HAP) to optically pure (S)‐1‐phenyl‐1,2‐ethanediol. Enzymatic assays revealed that YaCRI and YaCRII exhibited specific activities of 6·96 U mg−1 (99·8% e.e.) and 7·85 U mg−1 (99·9% e.e.), respectively, and showed moderate heat resistance at 40–50°C and acid tolerance at pH 5·0–6·0. An efficient whole‐cell two‐phase system was established using reductase‐expressing recombinant Escherichia coli. The conversion of 2‐HAP (20·0 g l−1) conversion with the solvent of dibutyl phthalate was approximately 70‐fold higher than in water. Furthermore, the two recombinant E. coli displayed biocatalyst activity and enantioselectivity towards several different carbonyl compounds, and E. coli BL21 (DE3)/pET‐28a‐yacrII showed a broad substrate spectrum.
Conclusions
A new whole‐cell recombinant E. coli‐based bioreduction system for enantiopure alcohol synthesis with high enantioselectivity at high substrate concentrations was developed.
Significance and Impact of the Study
We proposed a promising approach for the efficient preparation of enantiopure chiral alcohols.
Trans‐4‐Hydroxy‐l‐proline (trans‐Hyp) is a valuable chiral building block for the synthesis of pharmaceutical intermediates. Bioconversion of l‐proline using recombinant strain with ...proline‐4‐hydroxylase (P4H) is a preferred biocatalytic process in the economical production of trans‐Hyp. In this study, a recombinant E. coli overexpressing hydroxylase (P4H), γ‐glutamyl kinase and glutamate‐semialdehyde dehydrogenase (ProBA) genes were constructed by knocking out the key genes in the metabolism. These key genes contained putA encoding proline dehydrogenase (PutA) in the l‐proline metabolism and other catalytic enzyme genes, sucAB encoding α‐ketoglutarate dehydrogenase (SucAB), aceAK encoding isocitratelyase (AceA) and isocitrate dehydrogenase kinase/phosphatase (AceK) in the TCA cycle. This recombinant strain coupled the synthetic pathway of trans‐Hyp with TCA cycle of the host strain. It inhibited the consumption of l‐proline completely and promoted the accumulation of 2‐oxoglutarate (2‐OG) as a co‐substrate, which realized the highest conversion of glucose to trans‐Hyp. A fed‐batch strategy was designed, capable of producing 31·0 g l−1 trans‐Hyp from glucose. It provided a theoretical basis for commercial conversion of glucose to trans‐Hyp.
Significance and Impact of the Study:Trans‐4‐Hydroxy‐l‐proline (trans‐Hyp) is a valuable chiral building block for the synthesis of pharmaceutical intermediates. Unsatisfactory microbial bioconversion resulted in a low yield of trans‐Hyp. In this study, we blocked the unwanted blunting pathways of host strain and make the cell growth couple with the trans‐Hyp synthesis from glucose. Finally, a recombinant Escherichia coli with short‐cut and efficient trans‐Hyp biosynthetic pathway was obtained. It provided a theoretical basis for commercial production of trans‐Hyp.
Significance and Impact of the Study:Trans‐4‐Hydroxy‐l‐proline (trans‐Hyp) is a valuable chiral building block for the synthesis of pharmaceutical intermediates. Unsatisfactory microbial bioconversion resulted in a low yield of trans‐Hyp. In this study, we blocked the unwanted blunting pathways of host strain and make the cell growth couple with the trans‐Hyp synthesis from glucose. Finally, a recombinant Escherichia coli with short‐cut and efficient trans‐Hyp biosynthetic pathway was obtained. It provided a theoretical basis for commercial production of trans‐Hyp.
Herein is presented an efficient nonlinear filtering algorithm called the Gaussian-sum cubature Kalman filter (GSCKF) for the bearings-only tracking problem. It is developed based on the recently ...proposed cubature Kalman filter and is built within a Gaussian-sum framework. The new algorithm consists of a splitting and merging procedure when a high degree of nonlinearity is detected. Simulation results show that the proposed algorithm demonstrates comparable performance to the particle filter (PF) with significantly reduced computational cost.
BACKGROUND AND PURPOSE Dietary intake of ω‐3 polyunsaturated fatty acids (ω‐3 PUFAs) like eicosapentaenoic acid (EPA) decreases cancer risk, while arachidonic acid and other ω‐6 PUFAs increase risk, ...but the underlying mechanisms are unclear. Cytochrome P450 (CYP)‐derived epoxides contribute to enhanced tumourigenesis due to ω‐6 PUFA intake. Thus, ω‐6 arachidonic acid epoxides (EETs) inhibit apoptosis and stimulate proliferation by up‐regulating cyclin D1 expression in cells. The present study evaluated the corresponding ω‐3 PUFA epoxides and assessed their role in the regulation of cell proliferation.
EXPERIMENTAL APPROACH Four chemically stable EPA epoxides (formed at the 8,9‐, 11,12‐, 14,15‐ and 17,18‐olefinic bonds) were synthesized and tested against growth‐related signalling pathways in brain microvascular endothelial bEND.3 cells. Cell cycle distribution was determined by flow cytometry and cyclin gene expression by immunoblotting and real‐time PCR. The role of the p38 mitogen‐activated protein (MAP) kinase in cyclin D1 dysregulation was assessed using specific inhibitors and dominant‐negative expression plasmids.
KEY RESULTS The ω‐3 17,18‐epoxide of EPA decreased cell proliferation, interrupted the cell cycle in S‐phase and down‐regulated the cyclin D1/cyclin‐dependent kinase (CDK)‐4 complex, whereas the 8,9‐, 11,12‐ and 14,15‐epoxides were either inactive or enhanced proliferation. Cyclin D1 down‐regulation by 17,18‐epoxy‐EPA was mediated by activation of the growth‐suppressing p38 MAP kinase, but the alternate EPA‐epoxides were inactive.
CONCLUSIONS AND IMPLICATIONS The present findings suggest that the epoxide formed by CYP enzymes at the ω‐3 olefinic bond may contribute to the beneficial effects of ω‐3 PUFA by down‐regulating cyclin D1 and suppressing cell proliferation.
Breast cancer developed in familial BRCA1 mutation carriers bears striking similarities to sporadic basal-like breast tumors. The mechanism underlying the function of BRCA1 in suppressing basal-like ...breast cancer remains unclear. We previously reported that the deletion of p18(Ink4c) (p18), an inhibitor of G1 cyclin Ds-dependent CDK4 and CDK6, stimulates mammary luminal progenitor cell proliferation and leads to spontaneous luminal tumor development. We report here that germline mutation of Brca1 in p18-deficient mice blocks the increase of luminal progenitor cells, impairs luminal gene expression and promotes malignant transformation of mammary tumors. Instead of the luminal mammary tumors developed in p18 single-mutant mice, mammary tumors developed in the p18;Brca1 mice, similar to breast cancer developed in familial BRCA1 carriers, exhibited extensive basal-like features and lost the remaining wild-type allele of Brca1. These results reveal distinct functions of the RB and BRCA1 pathways in suppressing luminal and basal-like mammary tumors, respectively. These results also suggest a novel mechanism--causing luminal-to-basal transformation--for the development of basal-like breast cancer in familial BRCA1 carriers and establish a unique mouse model for developing therapeutic strategies to target both luminal and basal-like breast cancers.
Cell expansion is crucial for plant growth. It is well known that the phytohormone ethylene functions in plant development as a key modulator of cell expansion. However, the role of ethylene in the ...regulation of this process remains unclear. In this study, 2,189 ethylene-responsive transcripts were identified in rose (Rosa hybrida) petals using transcriptome sequencing and microarray analysis. Among these transcripts, an NAC (for no apical meristem NAM, Arabidopsis transcription activation factor ATAF, and cup-shaped cotyledon CUC)-domain transcription factor gene, RhNAC100, was rapidly and dramatically induced by ethylene in the petals. Interestingly, accumulation of the RhNAC100 transcript was modulated by ethylene via microRNA164-dependent posttranscriptional regulation. Overexpression of RhNAC100 in Arabidopsis (Arabidopsis thaliana) substantially reduced the petal size by repressing petal cell expansion. By contrast, silencing of RhNAC100 in rose petals using virus-induced gene silencing significantly increased petal size and promoted cell expansion in the petal abaxial subepidermis (P < 0.05). Expression analysis showed that 22 out of the 29 cell expansion-related genes tested exhibited changes in expression in RhNAC100-silenced rose petals. Moreover, of those genes, one cellulose synthase and two aquaporin genes (Rosa hybrida Cellulose Synthase2 and R. hybrida Plasma Membrane Intrinsic Protein1;1/2;1) were identified as targets of RhNAC100. Our results suggest that ethylene regulates cell expansion by fine-tuning the microRNA164/RhNAC100 module and also provide new insights into the function of NAC transcription factors.
Abstract
Polymer composites are gradually replacing traditional metal materials in the fields of aviation, aerospace, automotive and medicine due to their corrosion resistance, light weight and high ...strength. Moulding technology and organization morphology of polymer composite are key elements affecting the quality of products and their application, so a vacuum hot pressing process for graphene
x
/poly(ether ketone ketone) (PEKK) (x = 0%, 2%, 3%, 4%, 5%, 6%) composite powders is explored with particularly designed moulding parameters to achieve high conductive properties and good mechanical properties in graphene/PEKK composite sheet with thickness of 1.25 mm and diameter of 80 mm. The vacuum environment ensures that the graphene is not oxidized by air during hot pressing molding, which is essential for achieving conductive property in the graphene/PEKK composite; The hot pressing temperature of each graphene/PEKK composite powder is higher than glass transition temperature but lower than melting temperature, which ensures the graphene/PEKK composite powders is fully compacted and then graphene is fully lapped in the composite sheet. In addition, the graphene/PEKK composite sheet shows conductive property when the graphene content increases to 3wt%, and then the conductivity of the composites increases and then decreases with a peak value at 5wt% with increasing graphene content. By comparing the mechanical properties and microstructure morphology of the graphene/PEKK composite sheets, it was obtained that graphene content has an obvious effect on the mechanical properties of the composites, e.g., the mechanical properties will be increased as the graphene content increasing when graphene content is more than 3%. The graphene distribution law of the composite material with different graphene contents is analysed using a scanning electron microscope (SEM).
Long non-coding RNA (lncRNA) play important roles in many diseases. However, their roles in sporadic thoracic aortic aneurysm (STAA) are unclear. Therefore, the objective of this study was to ...construct an lncRNA–mRNA network and dissect lncRNAs that might contribute to the pathogenesis of STAA.
Differentially expressed lncRNAs and mRNAs between four ascending aortic specimens derived from STAA and four controls from patients who had undergone coronary artery bypass graft (CABG) were identified by microarray analysis. Gene Ontology (GO) enrichment and lncRNA–mRNA correlation analysis were implemented with differentially expressed genes. An lncRNA in the correlation network HOX transcript antisense intergenic RNA (HOTAIR) was selected as a candidate. HOTAIR expression was examined by quantitative real time polymerase chain reaction in STAA (n = 24) and controls (n = 24 CABG, n = 22; heart transplant donors, n = 2). HOTAIR expression was knocked down with siRNA in order to evaluate its role in apoptosis, cell proliferation, and expression of collagen types I and III.
Five percent of lncRNAs were significantly differentially expressed in STAA patient samples compared with controls. GO enrichment analysis suggested differentially expressed genes were significantly enriched in the process of extracellular matrix organisation and leukocyte migration. lncRNA–mRNA interaction network revealed HOTAIR was associated with genes involved in extracellular matrix organisation. Moreover, HOTAIR expression was significantly decreased in STAA specimens and it negatively correlated with aortic diameter. HOTAIR knockdown induced early and late apoptosis and reduced cell proliferation. Furthermore, both mRNA and protein levels of collagen types I and III expression were suppressed after HOTAIR knockdown.
Transcriptomic and lncRNA–mRNA correlation analysis revealed HOTAIR was downregulated in STAA and associated with genes involved in extracellular matrix remodelling. In vitro experiments confirmed that knockdown of HOTAIR could induce apoptosis and suppress collagen types I and III expression in human aortic smooth muscle cells.
Folate-dependent one-carbon cycle metabolism (FOCM) plays a critical role in maintaining genomic stability through regulating DNA biosynthesis, repair and methylation. Folate metabolites as well as ...other metabolites in the FOCM are hypothesized to be altered when cells transition from normal to cancerous state. Using cells at different stages in their development into colorectal cancer, the FOCM metabolites were profiled as an effort to phenotype the cells, and the metabolite levels were compared to the expressions of related genes. Here, we investigate whether there is a correlation between the metabolite levels, DNA methylation levels and the expression of the related genes that drive the levels of these metabolites. Using CRL1459, APC10.1, HCT116 and Caco-2, we show for the first time that FOCM metabolites correlate with the gene expression patterns. These differences follow a trend that may facilitate distinguishing colon cells at the different stages as they transition into cancerous state. The folate distribution and methionine levels were found to be key in determining the staging of the colon cells in CRC development. Also, expression of CBS, MTRR and MAT genes may facilitate distinguishing between untransformed and transformed colon cells.
This study aimed to analyse the existing evidence on the association between particulate matter 2.5 (PM2.5) and dementia, including two of its subtypes, namely, Alzheimer's disease (AD) and vascular ...dementia (VaD).
This was a systematic review and meta-analysis.
The PubMed, EMBASE, Cochrane and Web of Science databases were comprehensively searched for articles published between January 1900 and June 2022. All cohort studies that reported the influence of long-term exposure to PM2.5 on dementia, together with its subtypes, in adults aged ≥40 years, without any regional restriction were included. A random effects model was used to pool the hazard ratios (HRs) of PM2.5 for dementia, AD and VaD. Funnel plots, sensitivity analyses and subgroup analyses were performed to test publication bias and result stability. In addition, an explanation for the heterogeneity of the results was suggested.
In total, 20 articles were selected for review; 18 included results on the long-term effects of PM2.5 on dementia, 13 on AD, and eight on VaD. Three group meta-analyses were performed to obtain the HRs and 95% confidence intervals (CIs). The pooled HRs were 1.40 (95% CI 1.23, 1.60) for dementia, 1.47 (95% CI 1.22, 1.78) for AD and 2.00 (95% CI 1.30, 3.08) for VaD per 10.0 μg/m3 PM2.5 increase.
Long-term exposure to PM2.5 may increase the risk of dementia, including AD and VaD. These results highlight the need for further study on the detrimental impact of PM2.5 and the importance of strategies to mitigate increasing air pollution.
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