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In the Chinese strong-flavor baijiu (CSFB) fermentation ecosystem, the caproic acid-anabolism of caproic acid-producing bacteria (CPBs) is very important for improving the fermentation quality of CSFB. Therefore, it is necessary to thoroughly understand the types of CPBs and their caproic acid-anabolism characteristics. This minireview introduces readers to the diversity, phylogenetic relationship, physiological and metabolic characteristics, and caproic acid synthesis mechanism of CPBs isolated from the CSFB fermentation ecosystem as well as their synergistic metabolic relationships with other CPBs or non-CPBs. This paper provides a reference for understanding the in-situ caproic acid-anabolism pattern of CPBs from the CSFB fermentation ecosystem, and further provides a theoretical basis for the future targeted application of CPBs in CSFB fermentation and for CPBs culture engineering for the synthesis of high value-added caproic acid.

In order to improve the ability of a wild-type strain (BY-1) of Pichia pastoris to produce glutathione (GSH), a mutant strain designated BY-1-26 was obtained by atmospheric and room temperature plasma (ARTP) mutagenesis. To further improve the production of glutathione by the mutant during shake flask culture, 1,2,4-triazole was added to the culture medium. Finally, adaptive evolution of the mutant strain was carried out on a microbial microdroplet culture system (MMC) using 1,2,4-triazole as a screening factor. As a result, a high-yield glutathione-producing mutant strain BY-2-24 was obtained, and its genetic stability was investigated. The results showed that a high-yield glutathione-producing mutant strain could be obtained using the wild-type strain BY-1 by ARTP mutagenesis, primary resistance screening on gradient plates, adaptive evolution on MMC and shake-flask secondary screening. The yield of glutathione produced by BY-2-24 in shake flask culture was (312.13 ± 2.62) mg/L, which was 134.26% higher than that produced by the original strain, and this mutant still had good genetic stability after seven passages. Meanwhile, the biomass was increased by 118.33%, indicating that the growth ability of the mutant strain was improved compared to the original strain. The result of this study show that the combination of ARTP and MMC can be used as a simple and effective screening method for excellent mutant strains, which provides a reference for high throughput selection of target strains.

To investigate the mechanism of biofilm formation in Vibrio parahaemolyticus, its physiological characteristics were examined and the regulation of gene expression during biofilm formation was studied transcriptomic sequencing. Two strains of V. parahaemolyticus (ATCC 17802 and VP-0) that differ in their biofilms were evaluated for their physiological indexes such as extracellular polysaccharides, extracellular proteins, the signaling molecule autoinducer-2 (AI-2), cell permeability, and biofilm microscopic morphology, and the dynamic formation process of biofilm was investigated. The results showed that the biofilm formation process of the tested strains involved reversible adhesion (from 0 to 12 h), irreversible adhesion and micro-colony formation (from 12 to 48 h), maturation (from 48 to 72 h), and dissociation (from 72 to 144 h). The secretion rates of extracellular polysaccharides and proteins at the biofilm maturation stage of ATCC 17802 were 1.67 and 2.3 times higher than those of VP-0, respectively. The difference in signaling molecule content during biofilm formation was not significant (P > 0.05), and laser confocal scanning microscopy (LCSM) showed that ATCC 17802 had a greater degree of aggregation. Based on the results of transcriptomic sequencing, 802, 1 061, and 267 significantly differentially expressed genes were identified in three treatment groups (72 vs 8 h, 48 vs 8 h, and 72 vs 48 h), of which 506, 655 and 96 differentially expressed genes were down-regulated and 296, 406 and 171 differentially expressed genes were up-regulated, respectively. These differences were mainly involved in energy metabolism, flagellar system, transport system, and other aspects related to biofilm formation. The findings obtained in this study provide a theoretical basis for gene expression regulation during the dynamic formation of V. parahaemolyticus biofilm.

Water-dispersive Fe3O4@C nanoparticles have been successfully synthesized on a large scale via a one-pot template-free solvothermal process. The core–shell structure of the product was confirmed by transmission electron microscopy (TEM) observations. Fourier transform infrared (FT-IR) and X-ray photoelectron spectra (XPS) of the sample indicated that abundant amide carbonyl groups are formed on the surface of the Fe3O4@C nanoparticles. Based on the thermogravimetric analysis (TGA) results, the contents of Fe3O4 and carbon are calculated to be 77.5% and 6.1%, respectively. Magnetic measurements illustrated that the prepared composites are superparamagnetic magnets with a saturation magnetization of 69.2 emu g−1. Because of their hydrophilic core–shell structure, the prepared Fe3O4@C nanoparticles show a high bovine serum albumin (BSA) protein adsorption capacity (82.78 mg g−1) and a fast adsorption rate (60 min) in aqueous solutions. This work demonstrates that the prepared magnetic nanoparticles have promising applications in enzyme and protein immobilization. [ABSTRACT FROM AUTHOR]

Isocitrate dehydrogenase (IDH) family of proteins is classified into three subfamilies, namely, types I, II, and III. Although IDHs are widely distributed in bacteria, archaea, and eukaryotes, all type III IDHs reported to date are found only in prokaryotes. Herein, a novel type III IDH subfamily member from the marine microalga Phaeodactylum tricornutum (PtIDH2) was overexpressed, purified, and characterized in detail for the first time. Relatively few eukaryotic genomes encode this type of IDH and PtIDH2 shares the highest homology with marine bacterial monomeric IDHs, suggesting that PtIDH2 originated through a horizontal gene transfer event between a marine alga and a bacterium. Size-exclusion chromatography revealed that the native PtIDH2 is a homotetramer (∼320 kDa) in solution, comprising four monomeric IDH-like subunits (80 kDa each). Enzymatic characterization showed that PtIDH2 is a bivalent metal ion-dependent enzyme and Mn 2+ is the optimal activator. The recombinant PtIDH2 protein exhibited maximal activity at 35°C and pH 8.0 in the presence of Mn 2+ . Heat-inactivation analysis revealed that PtIDH2 is a cold-adapted enzyme. Kinetic analysis demonstrated that PtIDH2 is a completely NADP + -specific IDH with no detectable NAD + -associated catalytic activity. The three putative key NADP + -binding residues (His604, Arg615, and Arg664) in PtIDH2 were also evaluated by site-directed mutagenesis. The H 604 L/R 615 D/R 664 S triple mutant showed a 3.25-fold preference for NAD + over NADP + , implying that the coenzyme specificity of PtIDH2 can be converted from NADP + to NAD + through rational engineering approaches. Additionally, the roles of the conserved residues Ala718 and Leu742 in the thermostability of PtIDH2 were also explored by site-directed mutagenesis. We found that the L 742 F mutant displayed higher thermostability than wild-type PtIDH2. This study expands the phylogeny of the IDH family and provides new insights into the evolution of IDHs., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Huang, Zhao, Li, Wang, Xue and Zhu.)

The development of orally administrated heparin drugs requires a systematic understanding of the interaction between heparin and gut flora. The in vivo distribution of fluorescein-labeled heparin that is orally administrated by mice was observed using fluorescein microscopy. In addition, the stability of heparin in simulated gastric and intestinal fluids, as well as the in vitro degradation of heparin by gut flora were detected by HPLC. The results show that orally administrated heparin was mainly distributed in the gastrointestinal tract of mice, and exerted structural stability under the condition of simulated gastric and intestinal fluids in vitro. However, heparin could be degraded by intestinal flora cultured in medium containing heparin. In order to further study the effect of orally administrated heparin on intestinal flora in mice, the fecal microbiota 16S rRNA fragment of C57BL/6J mice was tested by the Illumina Mi-Seq high-throughput sequencing technology. Compared with the gut flora of mice that orally administrated by saline, the biodiversity of gut flora in mice with orally administrated heparin was decreased. The difference of microflora structure was not significant at the phylum level, and the relative abundance of Alistipes, Parasutterella and Akkermansia was increased at the genus level, and the relative abundance of Bilophila, Enterorhabdus, Ruminiclostridium, Prevotellaceae_UCG_001, Ruminiclostridium-9, Bacteroides, Lachnoclostridium, Candidatus, Saccharimonas, Intestinimonas and Dubosiella was reduced. These findings indicate that heparin could influence the gut flora of mice. In addition, no obvious toxic and side effects were found in mice that orally administrated heparin, suggesting the safety of orally administrated heparin.

In this study, we analyzed the kinetics of bioconversion of conjugated linoleic acid (CLA) by permeabilized Lactobacillus acidophilus cells. The effects of cell mass, linoleic acid (LA) concentration, reaction pH and temperature on the bioconversion of CLA by permeabilized cells were investigated and the model system of bioconversion of CLA was established. The results showed that the production of CLA was increased by permeabilized cells. The optimal cell mass, pH and temperature of bioconversion of CLA were 10 x 10(10) ufc/mL, 4.5 and 45 degrees C, respectively. A marked LA inhibition phenomenon existed, and the early reaction rate of producing CLA reached the maximum (17.8 microg/mL x min) when LA concentration was 0.6 mg/mL. Michaelis constant was obtained by double-reciprocal plot and Hanes-Woolf plot. The reaction rate equation followed the classic Michaelis-Mentent equation at the low LA concentration, while there was a marked LA inhibition phenomenon at the high LA concentration. With the evaluated model parameters, the model system appeared to provide a description for the bioconversion of CLA by permeabilized Lactobacillus acidophilus cells.