Chiral amino acids are extensively applied in the pharmaceutical, food, cosmetic, and agricultural industries. As a representative example, l -phosphinothricin ( l -PPT, a chiral non-natural amino acid) is a broad-spectrum herbicide. An NAD(H)-driven biocatalytic system for the asymmetric synthesis of chiral amino acids (focused on l -PPT) with high efficiency and low cost is highly desired. The key for the development of such biocatalytic system is to obtain an NADH-dependent biocatalyst with high catalytic performance toward l -PPT pro-ketone PPO. Herein, an engineered glutamate dehydrogenase from Lysinibacillus composti ( Lc GluDH) with desired activity was obtained by gene mining and protein engineering. In silico analyses suggested that the volume of substrate-binding pocket was substantially enlarged from 330.5 Å 3 to 409.6 Å 3 . The stability of Lc GluDH was increased ( T m value increased from 47.3 °C to 65.3 °C) by introducing positively charged amino acids or aromatic amino acids at position 375. The desired biocatalytic system was constructed by coupling the engineered Lc GluDH and an NAD + -dependent FDH. Through this biocatalytic system, the batch production of l -PPT exhibited high space-time yield (207.3 g ⋅ L −1 ⋅ day −1 ) with strict stereoselectivity ( ee of l -PPT>99%). Furthermore, eight other chiral amino acids were synthesised by the developed NAD(H)-driven biocatalytic system with high ee values.
MCT1 is highly expressed by activated and germinal center B cells. • Targeting MCT1 suppressed humoral response and autoimmunity. • Targeting MCT1 inhibited B cell survival by modulating NAD(H) redox state. Immunometabolism has garnered significant attention due to its essential roles in regulating B cell activation, proliferation and differentiation. Monocarboxylate transporters (MCTs), a family of solute carrier transporters responsible for lactate transport across biological membranes, have been shown to play a pivotal role in immune responses across various immune cell types. However, the expression and functional significance of MCTs in B cells remain poorly understood. In this study, we demonstrated that germinal center B cells and activated B cells exhibit elevated levels of MCT1 expression. Pharmacologically blocking MCT1 with AZD3965 inhibited both T-cell dependent and independent humoral response and ameliorated autoimmunity in the experimental autoimmune myasthenia gravis. Mechanically, MCT1 blockade led to the accumulation of intracellular lactate, which in turn suppressed NAD + regeneration, thereby inhibiting B cell activation and survival. Taken together, these findings suggest that MCT1 might be a potential therapeutical target in antibody-mediate autoimmune disease. Download: Download high-res image (95KB) Download: Download full-size image
Biohydrogen productions from xylose and glucose under dark condition were enhanced by the presence of natural Fe 3 O 4 . The electron equivalent of H 2 fractions accounted for 4.55 % and 5.69 % of the total given xylose and glucose in the experiments without Fe 3 O 4 , and that were correspondingly increased to 5.14 % and 6.50 % in the experiments with 100 mg/L of Fe 3 O 4 , respectively. Moreover, Fe 3 O 4 increased the total intracellular NAD(H) concentrations by 8.84 % and 8.37 %, and boosted the ratios of NADH/NAD + by 8.33 % and 17.72 % in xylose and glucose fermentation, respectively, comparing to the corresponding control experiments. The formation of electron couples of Fe(III)/Fe(II) during the iron oxide respiration and more generation of active extracellular polymeric substances components were determined as the important reasons for the improved biohydrogen production performance. Thus, a promotion mechanism of the internal “driving forces” from extracellular iron oxide respiration on the biohydrogen production was proposed.
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