Characterization of an Entner-Doudoroff pathway-activated Escherichia coli.

Background: Escherichia coli have both the Embden-Meyerhof-Parnas pathway (EMPP) and Entner-Doudoroff pathway (EDP) for glucose breakdown, while the EDP primarily remains inactive for glucose metabolism. However, EDP is a more favorable route than EMPP for the production of certain products.

Results: EDP was activated by deleting the pfkAB genes in conjunction with subsequent adaptive laboratory evolution (ALE).

Levulinic Acid-Inducible and Tunable Gene Expression System for .

AM1 is an efficient platform strain possessing biotechnological potential in formate- and methanol-based single carbon (C1) bioeconomy. Constitutive expression or costly chemical-inducible expression systems are not always desirable. Here, several glucose-, xylose-, and levulinic acid (LA)-inducible promoter systems were assessed for the induction of green fluorescent protein (GFP) as a reporter protein.

High-Throughput Screening of Acyl-CoA Thioesterase I Mutants Using a Fluid Array Platform.

Screening target microorganisms from a mutated recombinant library plays a crucial role in advancing synthetic biology and metabolic engineering. However, conventional screening tools have several limitations regarding throughput, cost, and labor. Here, we used the fluid array platform to conduct high-throughput screening (HTS) that identified 'TesA thioesterase mutants producing elevated yields of free fatty acids (FFAs) from a large (10) mutant library.

Oligo- and dsDNA-mediated genome editing using a tetA dual selection system in Escherichia coli.

The ability to precisely and seamlessly modify a target genome is needed for metabolic engineering and synthetic biology techniques aimed at creating potent biosystems. Herein, we report on a promising method in Escherichia coli that relies on the insertion of an optimized tetA dual selection cassette followed by replacement of the same cassette with short, single-stranded DNA (oligos) or long, double-stranded DNA and the isolation of recombinant strains by negative selection using NiCl2. This method could be rapidly and successfully used for genome engineering, including deletions, insertions, replacements, and point mutations, without inactivation of the methyl-directed mismatch repair (MMR) system and plasmid cloning.

Simultaneous utilization of glucose and xylose via novel mechanisms in engineered Escherichia coli.

After glucose, xylose is the most abundant sugar in lignocellulosic carbon sources. However, wild-type Escherichia coli is unable to simultaneously utilize both sugars due to carbon catabolite repression (CCR). In this paper, we describe GX50, an engineered strain capable of utilizing glucose and xylose simultaneously.

A simple and effective method for construction of Escherichia coli strains proficient for genome engineering.

Multiplex genome engineering is a standalone recombineering tool for large-scale programming and accelerated evolution of cells. However, this advanced genome engineering technique has been limited to use in selected bacterial strains. We developed a simple and effective strain-independent method for effective genome engineering in Escherichia coli.

Regulation of RCAN1 protein activity by Dyrk1A protein-mediated phosphorylation.

Two genes on chromosome 21, namely dual specificity tyrosine phosphorylation-regulated kinase 1A (Dyrk1A) and regulator of calcineurin 1 (RCAN1), have been implicated in some of the phenotypic characteristics of Down syndrome, including the early onset of Alzheimer disease. Although a link between Dyrk1A and RCAN1 and the nuclear factor of activated T cells (NFAT) pathway has been reported, it remains unclear whether Dyrk1A directly interacts with RCAN1. In the present study, Dyrk1A is shown to directly interact with and phosphorylate RCAN1 at Ser(112) and Thr(192) residues.

Dyrk1A-mediated phosphorylation of Presenilin 1: a functional link between Down syndrome and Alzheimer's disease.

The dual-specificity tyrosine(Y)-phosphorylation-regulated kinase 1A (Dyrk1A) gene is located on human chromosome 21 and encodes a proline-directed protein kinase that might be responsible for mental retardation and early onset of Alzheimer's disease (AD) in Down syndrome (DS) patients. Presenilin 1 (PS1) is a key component of the γ-secretase complex in the generation of β-amyloid (Aβ), an important trigger protein in the pathogenesis of AD. Increased Dyrk1A expression has been reported in human AD and DS brains.

In vitro development of a hemangioblast from a human embryonic stem cell, SNUhES#3.

Aims: Recent reports demonstrated that a hemangioblast population emerged during hematopoietic development in both mouse and human embryonic stem cell (hESC) differentiation cultures.

Main Methods: In this study, a new uncharacterized hESC line, SNUhES#3, was studied for its capacity to proliferate with STO cells and differentiate into hemangioblasts in co-culture with OP9 cells.

Key Findings: We were able to obtain CD34(+)CD45(-) cells from SNUhES#3 cells after 12 days of in vitro culture, and this cell population could be maximized to 12.

Capsaicin promotes the development of burst-forming units--erythroid (BFU-E) from mouse bone marrow cells.

Capsaicin, the pungent component of chilli peppers, is known to induce mediators of hematopoiesis. We investigated the effect of capsaicin on hematopoiesis in mouse progenitor cells. Treatment of mouse bone marrow cells with capsaicin induced the formation of colony of burst-forming units-erythroid (BFU-E).

COMP-Ang1: a designed angiopoietin-1 variant with nonleaky angiogenic activity.

Angiopoietin-1 (Ang1) has potential therapeutic applications in inducing angiogenesis, enhancing endothelial cell survival, and preventing vascular leakage. However, production of Ang1 is hindered by aggregation and insolubility resulting from disulfide-linked higher-order structures. Here, by replacing the N-terminal portion of Ang1 with the short coiled-coil domain of cartilage oligomeric matrix protein (COMP), we have generated a soluble, stable, and potent Ang1 variant, COMP-Ang1.

EphB ligand, ephrinB2, suppresses the VEGF- and angiopoietin 1-induced Ras/mitogen-activated protein kinase pathway in venous endothelial cells.

Interaction between ephrinB2 and EphB4 in endothelial cells at the arterial-venous capillary interface is critical for proper embryonic capillary morphogenesis. However, the intracellular downstream signaling of ephrinB2-EphB in vascular endothelial cells is unknown. This study examined the effect of ephrinB2-induced activation of EphB kinases on vascular endothelial growth factor (VEGF)- and angiopoietin-1 (Ang1)-induced Ras/mitogen-activated protein kinase (MAPK) signaling cascades in human umbilical vein endothelial cells (HUVECs).

Angiopoietin-1 negatively regulates expression and activity of tissue factor in endothelial cells.

Normally, tissue factor (TF) is not expressed on the surface of endothelial cells, but its expression can be induced by vascular endothelial growth factor (VEGF) and tumor necrosis factor (TNF)-a. However, the signaling pathway(s) affecting this induction is unknown. Using human umbilical vein endothelial cells, we found that inhibitors of guanine-cytosine-rich DNA binding protein and nuclear factor (NF)-kB suppressed VEGF- and TNF-a-induced expression and activity of TF.