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Asymmetric reduction of electronically activated alkenes by ene reductases (ERs) is an attractive approach for the production of enantiopure chiral products.

We aimed to investigate the effectiveness of endovascular therapy (EVT) versus intravenous thrombolysis (IVT) in patients with basilar artery occlusion (BAO), based on the information of advanced imaging.We analyzed data of stroke patients with radiologically confirmed BAO within 24 hours. BAO subjects were categorized into "top-of-the-basilar" syndrome (TOBS) and other types. An initial infarct size of70ml and a ratio of ischemic tissue to infarct volume of ≥1.8 was defined as "target mismatch." The primary outcome was a good outcome, defined as a modified Rankin Scale score of 0 to 3 at 3 months. Propensity score adjustment and inverse probability of treatment weighting (IPTW) propensity score methods were used.Among 474 BAO patients, 93 (19.6%) were treated with IVT prior to EVT, 91 (19.2%) were treated with IVT alone, 95 (20.0%) were treated with EVT alone, and 195 (41.1%) were treated with antithrombotic therapy. In IPTW analyses, we found no benefit of EVT over IVT for good outcome in either TOBS patients (odds ratio = 1.08, 95% confidence interval [CI] = 0.88-1.31) or those with other types (odds ratio = 1.13, 95% CI = 0.94-1.36). However, in patients with other types, if there existed a target mismatch, EVT was independently related to good outcome (odds ratio = 1.46, 95% CI = 1.17-1.81).The "target mismatch profile" seems to be a possible candidate selection standard of EVT for those with other types of BAO. Future studies should separate TOBS from other types of BAO, and try to use advanced imaging. ANN NEUROL 2022;92:97-106.

Background: D-xylonic acid is a versatile platform chemical with wide potential applications as the water reducer and disperser for cement and the precursor for 1,4-butanediol and 1,2,4-tributantriol. Microbial production of D-xylonic acid with bacteria like Gluconobacter oxydans from cheap lignocellulosic feedstock is generally regarded as one of the most promising and cost-effective methods to industrial production, but xylonic acid productivity is reduced by high substrate inhibition and hydrolysate inhibitors. Results: D-xylonic acid productivity of G. oxydans DSM2003 was improved by overexpressing the mGDH gene, which encoded the membrane-bound glucose dehydrogenase. Using the mutated plasmids based on pBBR1MCS-5 in our previous work, the recombinant strain G. oxydans/pBBR-R3510-mGDH with the significant improvement in D-xylonic acid production and the strengthened tolerance to hydrolysate inhibitors was obtained. The fed-batch biotransformation of D-xylose by this recombinant strain reached the record high titer (588.7 g/L), yield (99.4%), and space-time yield (8.72 g/L/h). Moreover, up to 246.4 g/L D-xylonic acid was produced directly from corn stover hydroxylates without detoxification at a yield of 98.9% and a space-time yield of 11.2 g/L/h. In addition, G. oxydans/pBBR-R3510-mGDH performed strong tolerance to typical inhibitors, i.e., formic acid, furfural, and 5-hydroxymethylfurfural. Conclusion: Through overexpression of mgdh in G. oxydans, we obtained a recombinant strain G. oxydans/pBBR-R3510-mGDH. It was capable to efficiently produce xylonic acid from the corn stover hydrolysate under high concentrations of inhibitors. The high D-xylonic acid productivity made G. oxydans/pBBR-R3510-mGDH an attractive choice for biotechnical production.

Objectives: 3,4-Dihydroxybutyric acid (3,4-DHBA) is a multi-functional C4 platform compound with wide applications in the synthesis of materials and pharmaceuticals. Currently, although the biosynthetic pathway for the production of 3,4-DHBA has been developed, low productivity still hampers its use on large scales. Here, a non-natural four-steps biosynthetic pathway was established in recombinant E. coli with a combinatorial strategy.Results: Firstly, several aldehyde dehydrogenases (ALDHs) were screened and characterized for catalyzing the dehydrogenation of 3,4-dihydroxybutanal (3,4-DHB) to 3,4-DHBA through in vitro enzyme assays. Secondly, a recombinant E. coli was successfully constructed to generate 3,4-DHBA from D-xylose by introducing the pathway containing BsGDH, YagF, PpMdlC and ALDH into E. coli with 3.04 g/L 3,4-DHBA obtained. Then, disruption of competing pathways by deleting xylA, ghrA, ghrB and adhP genes contributed to increase the accumulation of 3,4-DHBA by 87%. Final, fusion expression of PpMdlC and YagF resulted in an enhancement of 3,4-DHBA titer (7.71 g/L), as the highest titer reported so far.Conclusions: These results showed that deleting competing pathways and constructing fusion protein could significantly improve the 3,4-DHBA titer in engineered E. coli.

The purpose of this study was to investigate the relationship of methylenetetrahydrofolate reductase (MTHFR) gene C677T polymorphism and ischemic stroke Large-artery atherosclerosis (LAA) and Small-artery occlusion (SAO) subtypes in Chinese Han Population.One hundred and thirty LAA patients and one hundred and fifty-nine SAO patients data were prospectively collected. The control group was one hundred and ninety-eight subjects from the Medical Examination Center in the same period. Plasma homocysteine (Hcy) levels were determined by a double reagent enzymatic cycling method. The genotypes of MTHFR polymorphism (C677T) were determined using polymerase chain reaction-restriction fragment length polymorphism.As compared with the controls, hypertension prevalence, smoking consumption ratio were significantly higher, plasma Hcy levels increased significantly, plasma high-density lipoprotein cholesterol levels were significantly reduced in LAA and SAO groups (The present study suggests that hypertension, smoking, decreasing plasma high-density lipoprotein cholesterol levels and hyperhomocysteinemia (HHcy) are independent risk factors for LAA and SAO subtypes of ischemic stroke in Chinese Han Population. The MTHFR gene C677T C/T+T/T genotypes and T allele increases the risk of LAA and SAO subtypes of ischemic stroke.

L-erythrose, a rare aldotetrose, possesses various pharmacological activities. However, efficient L-erythrose production is challenging. Currently, L-erythrose is produced by a two-step fermentation process from erythritol. Here, we describe a novel strategy for the production of L-erythrose in Gluconobacter oxydans (G. oxydans) by localizing the assembly of L-ribose isomerase (L-RI) to membrane-bound sorbitol dehydrogenase (SDH) via the protein–peptide interactions of the PDZ domain and PDZ ligand. To demonstrate this self-assembly, green fluorescent protein (GFP) replaced L-RI and its movement to membrane-bound SDH was observed by fluorescence microscopy. The final L-erythrose production was improved to 23.5 g/L with the stepwise metabolic engineering of G. oxydans, which was 1.4-fold higher than that obtained using coexpression of SDH and L-RI in G. oxydans. This self-assembly strategy shows remarkable potential for further improvement of L-erythrose production.

2-keto-d-gluconic acid (2KGA) is widely used as a chemical intermediate in the cosmetic, pharmaceutical and environmental industries. Several microbial fermentation processes have been developed for production of 2KGA but these suffer from substrate/product inhibition, byproduct formation and low productivity. In previous work, we showed that 2KGA can be specifically produced from glucose (Glu) or gluconic acid (GA) by resting wild-type Gluconobacter oxydans DSM2003 cells, although substrate concentration was relatively low. In this study, we attempted to improve 2KGA productivity by G. oxydans DSM2003 by overexpressing the ga2dh gene, which encodes the membrane-bound gluconate-2-dehydrogenase enzyme (GA2DH). The ga2dh gene was overexpressed in G. oxydans DSM2003 under the control of three promoters, P tufB , P ga2dh or P ghp0169 , respectively. Among the recombinant strains obtained, G. oxydans_tufB_ga2dh showed a similar growth rate to that of the control strain and displayed the highest specific productivity of 2KGA from GA, which was increased nearly twofold compared with that of the control strain during batch biotransformation. When biocatalysis conditions were optimized, with provision of sufficient oxygen during biotransformation, up to 480 g/L GA was completely utilized over 45 h by resting cells of G. oxydans_tufB_ga2dh and 453.3 g/L 2KGA was produced. A productivity of 10.07 g/L/h and a yield of 95.3 % were obtained. Overexpression of the ga2dh gene also significantly improved the conversion of Glu to 2KGA. Under optimized conditions, 270 g/L Glu was converted to 321 g/L 2KGA over 18 h, with a yield of 99.1 % and a productivity of 17.83 g/L/h. The glucose concentrations during the batch biotransformation and the 2KGA productivities achieved in this study were relatively high compared with the results of previous studies. This study developed an efficient bacterial strain (G. oxydans_tufB_ga2dh) for the production of 2KGA by overexpressing the ga2dh gene in G. oxydans. Supply of sufficient oxygen enhanced the positive effect of gene overexpression on 2KGA production. Gluconobacter oxydans_tufB_ga2dh is thus a competitive species for use in 2KGA production.

Membrane-bound alcohol dehydrogenase (mADH) was overexpressed in Gluconobacter oxydans DSM 2003, and the effects on cell growth and glycolic acid production were investigated. The transcription levels of two terminal ubiquinol oxidases (bo3 and bd) in the respiratory chain of the engineered strain G. oxydans-adhABS were up-regulated by 13.4- and 3.8-fold, respectively, which effectively enhanced the oxygen uptake rate, resulting in higher resistance to acid. The cell biomass of G. oxydans-adhABS could increase by 26%-33% when cultivated in a 7L bioreactor. The activities of other major membrane-bound dehydrogenases were also increased to some extent, particularly membrane-bound aldehyde dehydrogenase (mALDH), which is involved in the catalytic oxidation of aldehydes to the corresponding acids and was 1.26-fold higher. Relying on the advantages of the above, G. oxydans-adhABS could produce 73.3gl-1 glycolic acid after 45h of bioconversion with resting cells, with a molar yield 93.5% and a space-time yield of 1.63gl-1h-1. Glycolic acid production could be further improved by fed-batch fermentation. After 45h of culture, 113.8gl-1 glycolic acid was accumulated, with a molar yield of 92.9% and a space-time yield of 2.53gl-1h-1, which is the highest reported glycolic acid yield to date.