Ectoine is an amino acid derivative and an important natural product in halophilic microorganisms. It plays an important role in protecting cells and stabilizing biological macromolecules, and can be widely used in biomedical fields such as drug preparation adjuvants, organ transplantation and preservation, skin wound repair and cosmetics. Due to the medical value and commercial market demand of ectoine, this article summarized the recent advances in the microbial production of ectoine, including the mutation and breeding of hyper-producing strains, construction of genetically and metabolically engineered strains, optimization of fermentation processes, and extraction and purification processes. The application of multi-omics technologies and computational biology to develop an ectoine producing cell factory was prospected, with the aim to provide a reference for ectoine overproduction.
Amino Acids, Diamino/metabolism* ; Fermentation

Oxidosqualene cyclases(OSCs), belonging to a multigene family, can convert a common precursor 2,3-oxidosqualene into various types of triterpene skeletons. In this study, primers were designed according to the analysis of Siraitia grosvenorii transcriptome data, and two OSC genes SgAS1(GenBank No. QDO67189.1) and SgAS2(GenBank No. QDO67190.1) were cloned. The open reading frame(ORF) of SgAS1 was 2 262 bp, encoding 754 amino acids, and the ORF of SgAS2 was 2 289 bp, encoding 762 amino acids. Real-time quantitative PCR results demonstrated that the two SgOSCs genes showed different expression patterns in stems, leaves, and different stages of fruits. Phylogenetic analysis showed that both SgAS1 and SgAS2 were clustered with β-amyrin synthases into a branch, but further functional characterization using yeast heterologous expression found that SgAS1 was inactive and SgAS2 could produce β-amyrin as the sole product. Multiple sequence alignments revealed that SgAS2 had a conserved MWCYCR sequence related to β-amyrin biosynthesis, while SgAS1 had an unusual LFCYTR sequence, for which the authors performed site-directed mutagenesis analysis of this sequence and found that tryptophan residue(W) was the key amino acid residue that affected the function of SgOSCs. In addition, the authors transformed the monofunctional β-amyrin synthase SgAS2 into the chassis strain GH1, which was previously modified by the research group, and increased the yield of β-amyrin to 44.05 mg·L~(-1). This study first reported the monofunctional β-amyrin synthase SgAS2 from S. grosvenorii and conducted site-directed mutagenesis and synthetic biology investigation on it, providing a valuable resource for the directed biosynthesis of triterpenoids.
Phylogeny ; Triterpenes/metabolism* ; Cloning, Molecular ; Amino Acids

The synthesis of fine chemicals using multi-enzyme cascade reactions is a recent hot research topic in the field of biocatalysis. The traditional chemical synthesis methods were replaced by constructing in vitro multi-enzyme cascades, then the green synthesis of a variety of bifunctional chemicals can be achieved. This article summarizes the construction strategies of different types of multi-enzyme cascade reactions and their characteristics. In addition, the general methods for recruiting enzymes used in cascade reactions, as well as the regeneration of coenzyme such as NAD(P)H or ATP and their application in multi-enzyme cascade reactions are summarized. Finally, we illustrate the application of multi-enzyme cascades in the synthesis of six bifunctional chemicals, including ω-amino fatty acids, alkyl lactams, α, ω-dicarboxylic acids, α, ω-diamines, α, ω-diols, and ω-amino alcohols.
Amino Acids ; Biocatalysis ; Amino Alcohols ; Coenzymes/metabolism* ; Diamines

Taking the typical metabolic control product-guanosine as an example, the method of metabolic flux shift investigation based on process multi-levels parameter correlation analysis was established. The metabolic pathway, multi-parameter correlation, accumulation of amino acid and organic acid during guanosine fermentation process were integratively analyzed. The metabolic flux shift from HMP to EMP was ascertained, which was assumed to be caused by the accumulation of ammonium ion. The subsequent optimization based on controlling flux distribution between EMP and HMP did improve the yield by 35% when the metabolic flux shift was prevented.
Amino Acids ; metabolism ; Ammonia ; metabolism ; Fermentation ; Guanosine ; metabolism

Amino acids are essential nutrients for humans and have a wide range of biological functions. They are the constituent units of protein and energy metabolites. In addition, they are also widely involved in the maintenance and regulation of various physiological functions, and play a role in transcription, translation, post-translational modification and other levels. The liver is a key metabolic organ, and it acts as a hub that connects the metabolism of various tissues. Amino acid sensing plays a very important role in the regulation of hepatic glucose and lipid metabolism. Therefore, accurately sensing the levels of intracellular and extracellular amino acids is the key to maintaining cell homeostasis. There are several well-known amino acid sensors in eukaryotic cells, such as general control non-derepressible-2 (GCN2), mammalian target of rapamycin (mTOR) and taste receptors, which play an important role in maintaining metabolic homeostasis. This article gives a detailed introduction to the role and mechanism of amino acids in regulating hepatic glucose and lipid metabolism, laying a foundation for further exploration of amino acid sensing mechanism and treatment of hepatic glucose and lipid metabolism disorders.
Amino Acids ; Glucose/metabolism* ; Homeostasis ; Humans ; Lipid Metabolism ; Liver

The isolated dog brain was perfused for a period of 4 minutes with blood normal in all constituents, but pathologically low in oxygen in order to follow the metabolic response of the brain to anoxia. During anoxic perfusion, the brain appears to subsist on the free amino acids in the brain and on glucose taken up from the perfusion blood. Oxygen uptake is relatively constant increasing temporarily immediately after anoxia. The lactic acid formed within the brain during anoxia is not released in any quantity; instead it appears to be metabolized in the brain following the period of anoxia causing a lower than norma1 uptake of glucose. Brain ATP and GrP levels decrease significantly, but not markedly indicating that the brain's energy requirements are being partially met during anoxic perfusion.
Amino Acids/*metabolism ; Animals ; Anoxia/*metabolism ; Brain/*metabolism ; Dogs ; Electroencephalography ; Glucose/*metabolism ; Lactates/*metabolism ; Oxygen Consumption

This study aimed to examine the effect of the 24 N-terminal amino acids (N24) of p55PIK, a regulatory subunit of phosphatidylinositol 3-kinase (PI3K), on the endotoxin lipopolysaccharide (LPS)-stimulated release of the cytokines (CKs) by HaCaT cells. The fusion protein, trans-acting activator of transcription (TAT)-N24 (an experimental peptide, EP) containing the N24 of PI3K-p55PIK, was constructed, and TAT-N24 fusion peptide was expressed and identified in BL21 E·coli. HaCaT cells (a human keratinocyte cell line) was cultured and stimulated by LPS at 100 ng/mL for 1, 2, 4, 8, 16 or 24 h, or by LPS at 10, 100 ng/mL, 1, 10 or 100 μg/mL of for 4 h. Changes in the protein and mRNA levels of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and interleukin-8 (IL-8) released by HaCaT cells following EP intervention were determined by enzyme-linked immunosorbent assay (ELISA) and real-time polymerase chain reaction (PCR). Immunofluorescence confocal laser scanning microscopy was utilized to detect the protein expression and translocation of the p65 subunit of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB p65) in HaCaT cells. The expression of the NF-κB inhibitor alpha (IκB-α) protein in LPS-stimulated HaCaT cells after the EP intervention was measured by Western blotting. The results showed that EP treatment increased TNF-α secretion from HaCaT cells. EP at certain concentrations could effectively inhibit the LPS-stimulated release of TNF-α, IL-6 and IL-8 from HaCaT cells. The ELISA assay demonstrated that the concentrations of TNF-α, IL-6 and IL-8 in the supernatants of LPS-stimulated cells were reduced from 208.06±30.18, 86.4±9.78 and 260.59±54.05 pg/mL to 121.78±22.26, 53.18±7.36 and 125.08±35.17 pg/mL, respectively, in the supernatants of cells treated by LPS and EP combined. Real-time PCR also revealed that the expression of the three pro-inflammatory CKs was significantly decreased after EP intervention. Immunofluorescence confocal laser scanning microscopy showed that NF-κB p65 protein was primarily expressed in the cytoplasm of non-stimulated HaCaT cells. After LPS stimulation, NF-κB p65 was translocated into the nucleus, and the nuclear expression of this protein increased. The nuclear NF-κB p65 protein expression was inhibited after the addition of EP. Western blotting showed that IκB-α expression began to decrease 30 min after LPS stimulation and declined to a trough 4 h later. IκB-α expression began to gradually recover 16 h after LPS stimulation but remained at a lower-than-normal level at 24 h. Greater IκB-α expression was found in cells treated with LPS and EP combined than those treated with LPS alone. It was concluded that EP can effectively inhibit the LPS-stimulated expression of TNF-α, IL-6, and IL-8, which involves the inhibition of the hydrolysis of IκB-α and thereby blockage of the nuclear translocation of NF-κB p65.
Amino Acids ; metabolism ; Cell Line ; Cytokines ; metabolism ; Endotoxins ; metabolism ; Humans ; Inflammation ; metabolism ; Phosphatidylinositol 3-Kinases ; metabolism

Cyclodipeptide (CDP) composed of two amino acids is the simplest cyclic peptide. These two amino acids form a typical diketopiperazine (DKP) ring by linking each other with peptide bonds. This characteristic stable ring skeleton is the foundation of CDP to display extensive and excellent bioactivities, which is beneficial for CDPs' pharmaceutical research and development. The natural CDP products are well isolated from actinomycetes. These bacteria can synthesize DKP backbones with nonribosomal peptide synthetase (NRPS) or cyclodipeptide synthase (CDPS). Moreover, actinomycetes could produce a variety of CDPs through different enzymatic modification. The presence of these abundant and diversified catalysis indicates that actinomycetes are promising microbial resource for exploring CDPs. This review summarized the pathways for DKP backbones biosynthesis and their post-modification mechanism in actinomycetes. The aim of this review was to accelerate the genome mining of CDPs and their isolation, purification and structure identification, and to facilitate revealing the biosynthesis mechanism of novel CDPs as well as their synthetic biology design.
Actinobacteria/metabolism* ; Actinomyces/metabolism* ; Biological Products/metabolism* ; Bacteria/metabolism* ; Diketopiperazines/metabolism* ; Amino Acids

In the 1920's, Warburg reported an observation that cancer cells depend on glycolysis even in the presence of available oxygen likely due to impaired function of mitochondria. Since then, this Warburg s effect has been the most important hypothesis in cancer metabolism and is considered as a seventh hallmark of many human cancers. Aerobic glycolysis was originally attributable to increased bioenergetic needs in rapidly proliferating cancer cells. Recently, biosynthetic aspects of aerobic glycolysis, which reprograms cancer metabolism to synthesize macromolecules such as nucleotides, fatty acids, amino acids, etc., are under active investigation. Introduction of positron emission tomography (PET) and metabolic radiotracers including F-18 flurorodeoxyglucose (FDG) and C-11 acetate made it possible to image cancer metabolism in vivo and to renew the interests on this issue. Studies have found that cancer cells with highly glycolysis features are associated with resistance to many chemotherapeutic regimens and radiation treatment. Therefore, development of glycolytic inhibitors can have an incremental effect to conventional treatments. In addition, functional imaging with metabolic radiotracers will continuously play important roles in detecting cancers and monitoring therapeutic responses to novel anti-metabolic approaches to cancer cells.
Amino Acids ; Electrons ; Energy Metabolism ; Fatty Acids ; Glycolysis ; Humans ; Mitochondria ; Nucleotides ; Oxygen ; Positron-Emission Tomography

OBJECTIVETo study the changes of soluble saccharides and amino acids during the growing period of Coptis chinensis.

METHODAmino acid contents were estimated by Indene tri-ketone colorimetric method; saccharide contents were determined by anthracene ketone method.

RESULT AND CONCLUSIONThe contents of soluble saccharides and amino acids in different organs were low in plants of 1-2 years old. And they were increased in plants of 3-4 years old. At maturation (5 years), the contents of amino acids kept on increasing in rhizome and flower stalk, while decreased in fibre and leaves, and the contents of soluble saccharides showed the downtrend in rhizome, leaves and fibre, except the uptrend in flower stalk.

Amino Acids ; metabolism ; Coptis ; growth & development ; metabolism ; Drugs, Chinese Herbal ; metabolism ; Polysaccharides ; metabolism ; Solubility ; Time Factors