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

Bacillus alcalophilus DTY1, one moderate halophytic bacterium isolated from saline soil in Loess Plateau of China, was characterized with efficient production of ectoine. In this study, the gene cluster ectABC taking in charge of biosynthesizing ectoine was cloned from the genomic library of strain DTY1. Nucleotide sequencing indicated that ectA, ectB and ectC were predicted to encode peptides of 169, 428 and 132 amino acids, respectively. The deduced amino acid sequences of EctA, EctB and EctC share 59%, 81% and 81% identity to 2,4-diaminobutyric acid acetyltransferase, 2,4-diaminobutyric acid transaminase and ectoine synthase of B. halodurans C-125, respectively. A fragment containing ectABC genes was introduced into B. cereus Z, which made the transgenic Z cells increased tolerance to salt, remarkably. HPLC analysis of ectoine in the transgenic Z cells revealed that 70.1 mg/g ectoine was detected in 1.0% NaCl medium and 118.6 mg/g ectoine in 5.0% NaCl medium. Furthermore, as the concentration of salt increased, transgenic Z cells accumulated more ectoine. These results suggest that ectoine is an important facet in B. alcalophilus DTY1 to high-osmolarity surroundings, and the expression of ectABC is induced by salt strength.
Amino Acid Sequence ; Amino Acids, Diamino ; biosynthesis ; genetics ; physiology ; Bacillus ; classification ; genetics ; Bacillus cereus ; genetics ; metabolism ; Bacterial Proteins ; genetics ; metabolism ; Base Sequence ; Cloning, Molecular ; Gene Expression Regulation, Bacterial ; Genes, Bacterial ; genetics ; Molecular Sequence Data ; Osmotic Pressure ; Sodium Chloride ; metabolism ; pharmacology

This study is to investigate the effects of fluvastatin on the activation of p38 mitogen-activated protein kinase (p38 MAPK) and cAMP response element-binding protein (CREB1) in glomerular mesangial cells under high concentration of glucose. High concentration glucose and fluvastatin were used to stimulate the cultured rat glomerular mesangial cells (GMCs) in vitro. The protein expressions of p38 MAPK, CREB1, p-p38 MAPK and p-CREB1 were observed with Western blotting. TGF-beta1 and fibronectin (FN) mRNA were measured with reverse transcription and polymerase chain reaction (RT-PCR). The protein synthesis of laminine (LN) and type IV collagen in the supernatants of the GMCs were detected with radioimmunoassay. Compared with low glucose control group, the expressions of p-p38 MAPK, p-CREB1 were increased obviously in high glucose group, TGF-beta1 mRNA and FN mRNA, LN and type IV collagen in the supernatants were increased significantly in GMCs under high concentration glucose medium. The expression levels of p-p38 MAPK, p-CREB1, TGF-beta1 mRNA, and FN mRNA, LN and type IV collagen in the supernatants were significantly lower in the fluvastatin group than those in the high concentration glucose group. It is concluded that fluvastatin can inhibit over production of TGF-beta1 and ECM proteins in GMCs under high concentration of glucose, partly by regulating the phosphorylation of p38 MAPK and CREB1.
Amino Acids, Diamino ; metabolism ; Animals ; Cell Proliferation ; Cells, Cultured ; Collagen Type IV ; metabolism ; Cyclic AMP Response Element-Binding Protein ; metabolism ; Fatty Acids, Monounsaturated ; pharmacology ; Fibronectins ; genetics ; metabolism ; Glucose ; administration & dosage ; Hydroxymethylglutaryl-CoA Reductase Inhibitors ; pharmacology ; Indoles ; pharmacology ; Male ; Mesangial Cells ; cytology ; metabolism ; Phosphorylation ; RNA, Messenger ; metabolism ; Rats ; Rats, Wistar ; Transforming Growth Factor beta1 ; genetics ; metabolism ; p38 Mitogen-Activated Protein Kinases ; metabolism

The adaptor protein, LAD/TSAd, plays essential roles in T cell activation. To further understand the functions of this protein, we performed yeast two-hybrid screening using TSAd as bait and identified 67 kDa laminin binding protein (LBP) as the interacting partner. Subsequently, TSAd-LBP interaction was confirmed in D1.1 T cell line. Upon costimulation by T cell receptor (TCR) plus laminin crosslinking or TCR plus integrin alpha6 crosslinking, LBP was coimmunoprecipitated with TSAd. Moreover, TCR plus laminin costimulation-dependent T cell migration was enhanced in D1.1 T cells overexpressing TSAd but was disrupted in D1.1 cells overexpressing dominant negative form of TSAd or TSAd shRNA. These data show that, upon TCR plus integrin costimulation, TSAd associates with LBP and mediates T lymphocyte migration.
Adaptor Proteins, Signal Transducing/genetics/*metabolism ; Amino Acids, Diamino/metabolism ; Carrier Proteins/*metabolism ; *Cell Movement ; Humans ; Integrin alpha6/metabolism ; Jurkat Cells ; Mutation ; Protein Binding ; RNA, Small Interfering/genetics ; *Receptor Cross-Talk ; Receptors, Antigen, T-Cell/metabolism ; T-Lymphocytes/*metabolism ; Transgenes ; Two-Hybrid System Techniques

OBJECTIVETo study the anti-inflammatory and analgesic activities of diethyl 1,3-dicyclohexyl-1,2,3,6-tetrahydropyrimidine-4,5-dicarboxylate (ZL-5010) in vivo and in vitro.

METHODSThe analgesic effect of ZL-5010 was evaluated by acetic acid-induced writhing response in mice, and the anti-inflammatory effects was assessed in mice with xylene-induced ear edema and in rats with carrageenan-induced paw edema. Mouse peritoneal exudate cells activated by bacterial lipopolysaccharides (LPS) were used to evaluate the anti-inflammatory effect of ZL-5010 in vitro. The levels of interleukin-1β (IL -1β) and tumor necrosis factor-α (TNF-α) in the cell culture supernatant were measured using enzyme-linked immunosorbent assay (ELISA).

RESULTSAt the doses of 0.25 and 0.5 mmol/kg, ZL-5010 administered by gavage once daily for 3 days significantly reduced acetic acid-induced writhing frequency and suppressed xylene-induced ear edema in mice, and alleviated paw edema induced by carrageenan in rats (P<0.05). The agent also inhibited the production of the pro-inflammatory cytokines IL-1β and TNF-α by LPS-induced mouse peritoneal exudate cells in vitro, with the statistically significant minimum effective concentrations of 10 and 20 µmol/L, respectively (P<0.05).

CONCLUSIONZL-5010 administered by gavage has anti-inflammatory and analgesic effects in mice and rats, and in mouse peritoneal exudate cell cultures, the agent also inhibits the production of the pro-inflammatory cytokines IL-1β and TNF-α.

Amino Acids, Diamino ; pharmacology ; therapeutic use ; Analgesics ; pharmacology ; therapeutic use ; Animals ; Anti-Inflammatory Agents ; pharmacology ; therapeutic use ; Cyclohexanes ; pharmacology ; therapeutic use ; Female ; Interleukin-1beta ; metabolism ; Male ; Mice ; Mice, Inbred Strains ; Pyrimidines ; pharmacology ; therapeutic use ; Rats ; Rats, Sprague-Dawley ; Tumor Necrosis Factor-alpha ; metabolism