• Current Research on Agriculture and Life Sciences
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• v.32 no.4
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• pp.199-202
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• 2014

The bioconversion of cellulosic biomass hydrolyzates consisting mainly of glucose and xylose requires the use of engineered Saccharomyces cerevisiae expressing a heterologous xylose pathway. However, there is concern that a fungal xylose pathway consisting of NADPH-specific xylose reductase (XR) and $NAD^+$-specific xylitol dehydrogenase (XDH) may result in a cellular redox imbalance. However, the glycerol biosynthesis and glycerol degradation pathways of S. cerevisiae, termed here as the glycerol cycle, has the potential to balance the cofactor requirements for xylose metabolism, as it produces NADPH by consuming NADH at the expense of one mole of ATP. Therefore, this study tested if the glycerol cycle could improve the xylose metabolism of engineered S. cerevisiae by cofactor balancing, as predicted by an in-silico analysis using elementary flux mode (EFM). When the GPD1 gene, the first step of the glycerol cycle, was overexpressed in the XR/XDH-expressing S. cerevisiae, the glycerol production significantly increased, while the xylitol and ethanol yields became negligible. The reduced xylitol yield suggests that enough $NAD^+$ was supplied for XDH by the glycerol cycle. However, the GPD1 overexpression completely shifted the carbon flux from ethanol to glycerol. Thus, moderate expression of GPD1 may be necessary to achieve improved ethanol production through the cofactor balancing.

• Journal of Microbiology and Biotechnology
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• v.30 no.2
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• pp.271-278
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• 2020

Glycerol dehydrogenase (GlyDH) catalyzes the oxidation of glycerol to dihydroxyacetone (DHA), which is the first step in the glycerol metabolism pathway. GlyDH has attracted great interest for its potential industrial applications, since DHA is a precursor for the synthesis of many commercially valuable chemicals and various drugs. In this study, GlyDH from Klebsiella pneumoniae (KpGlyDH) was overexpressed in E. coli and purified to homogeneity for biochemical and molecular characterization. KpGlyDH exhibits an exclusive preference for NAD⁺ over NADP⁺. The enzymatic activity of KpGlyDH is maximal at pH 8.6 and pH 10.0. Of the three common polyol substrates, KpGlyDH showed the highest k_cat/K_m value for glycerol, which is three times higher than for racemic 2,3-butanediol and 32 times higher than for ethylene glycol. The k_cat value for glycerol oxidation is notably high at 87.1 ± 11.3 sec^-1. KpGlyDH was shown to exist in an equilibrium between two different oligomeric states, octamer and hexadecamer, by size-exclusion chromatography analysis. KpGlyDH is structurally thermostable, with a T_m of 83.4℃, in thermal denaturation experiment using circular dichroism spectroscopy. The biochemical and biophysical characteristics of KpGlyDH revealed in this study should provide the basis for future research on its glycerol metabolism and possible use in industrial applications.

• The Plant Pathology Journal
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• v.37 no.1
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• pp.36-46
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• 2021

Acidovorax citrulli (Ac) is the causal agent of bacterial fruit blotch (BFB) in watermelon, a disease that poses a serious threat to watermelon production. Because of the lack of resistant cultivars against BFB, virulence factors or mechanisms need to be elucidated to control the disease. Glycerol-3-phosphate dehydrogenase is the enzyme involved in glycerol production from glucose during glycolysis. In this study, we report the functions of a putative glycerol-3-phosphate dehydrogenase in Ac (GlpdAc) using comparative proteomic analysis and phenotypic observation. A glpdAc knockout mutant, AcΔglpdAc(EV), lost virulence against watermelon in two pathogenicity tests. The putative 3D structure and amino acid sequence of GlpdAc showed high similarity with glycerol-3-phosphate dehydrogenases from other bacteria. Comparative proteomic analysis revealed that many proteins related to various metabolic pathways, including carbohydrate metabolism, were affected by GlpdAc. Although AcΔglpdAc(EV) could not use glucose as a sole carbon source, it showed growth in the presence of glycerol, indicating that GlpdAc is involved in glycolysis. AcΔglpdAc(EV) also displayed higher cell-to-cell aggregation than the wild-type bacteria, and tolerance to osmotic stress and ciprofloxacin was reduced and enhanced in the mutant, respectively. These results indicate that GlpdAc is involved in glycerol metabolism and other mechanisms, including virulence, demonstrating that the protein has pleiotropic effects. Our study expands the understanding of the functions of proteins associated with virulence in Ac.

• Applied Biological Chemistry
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• v.51 no.1
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• pp.60-64
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• 2008

The aerial parts of Spathiphyllum cannifolium were extracted with 80% aqueous MeOH and the concentrated extract was partitioned with EtOAc, n-BuOH and $H_2O$, successively. From the EtOAc fraction, three compounds were isolated through the repeated silica gel, ODS, Sephadex LH-20 column chromatographies. From the results of physico-chemical data including NMR, MS and IR, the chemical structures of the compounds were determined as stigmasterol (1), $(2S)1-O-linolenoyl-2-0-myristoyl-3-0-{\beta}-D-galactopyranosyl-sn-glycerol$ (2) and stigmasterol glucoside (3). They were the first to be isolated from Spathiphyllum cannifolium.

• Food Science and Biotechnology
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• v.18 no.1
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• pp.95-102
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• 2009

The aerial parts of garland (Chrysanthemum coronarium L.) were extracted in 80% aqueous methanol (MeOH) and the concentrated extract was then partitioned using ethyl acetate (EtOAc), n-butanol (n-BuOH), and $H_2O$, successively. EtOAc and n-BuOH fractions resulted in 4 glycerides with the application of octadecyl silica gel and silica gel column chromatography. The chemical structures of the glycerides were determined using several spectroscopic methods, including nuclear magnetic resonance (NMR) and mass spectrometry (MS) as (2S)-1-O-palmitoyl-sn-glycerol (1), (2S)-1-O-oleoyl-2-O-oleoyl- 3-O-$\beta$-D-galactopyranosyl-sn-glycerol (2), (2S)-1-O-palmitoyl-2-O-linoleoyl-3-O-phosphorouscholine-sn-glycerol (3), and (2S)-1-O-linolenoyl-2-O-palmitoyl-3-O-[$\alpha$-D-galactopyrasyl-($1{\rightarrow}6$)-$\beta$-D-galactopyranosyl]-sn-glycerol (4). The free fatty acids of these glycerides were determined with gas chromatography (GC)-MS analysis following alkaline hydrolysis and methylation. These glycerides demonstrated an inhibitory effect on acyl-CoA: cholesterol acyltransferase (ACAT, compound 1: $45.6{\pm}0.2%$ at $100{\mu}g/mL$), diacylglycerol acyltransferase (DGAT, compound 1: $59.1{\pm}0.1%$ at $25{\mu}g/mL$), farnesyl protein transferase (FPTase, compound 2: $98.0{\pm}0.1%$; compound 3: $55.2{\pm}0.1%$ at $100{\mu}g/mL$), and $\beta$-secretase ($IC_{50}$, compound 4: $2.6{\mu}g/mL$) activity. This paper is the first report on the isolation of these glycerides from garland and their inhibitory activity on ACAT, DGAT, FPTase, and $\beta$-secretase.

• Journal of the Korean Chemical Society
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• v.68 no.1
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• pp.32-39
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• 2024

Glycerol dehydrogenase (GlyDH) plays a crucial role in the glycerol metabolism pathway by catalyzing the oxidation of glycerol to dihydroxyacetone (DHA). Previous studies of GlyDH have predominantly focused on unraveling the structural features of the active site and its binding interactions with ligand. However, the structural details of GlyDH in complex with both NAD⁺ and the substrate bound have remained elusive. In this study, we present the crystal structures of Klebsiella pneumoniae GlyDH (KpGlyDH) in the absence and presence of NAD⁺ at a resolution of 2.1 Å. Notably, both structures reveal the binding of the substrate, ethylene glycol, to the zinc ion. Interestingly, a significant change in the coordination number of the zinc ion is observed, with three in the absence of NAD⁺ and four in its presence. These findings shed light on the structural aspects of GlyDH and its interactions with NAD⁺ and the substrate.

• Proceedings of the Korean Society of Applied Pharmacology
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• 2003.11a
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• pp.111-111
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• 2003

Green tea have been widely reported as functional foods because of their various bioactivities. In the present study, we used 3T3-Ll cells model of white adipocytes to clarify whether green tea and its main pharmaceutically effective compounds (EGCG, caffeine and theanine) prevent obesity. Cellular viability, glycerol-3-phosphate dehydrogenase activity, glycerol release and HSL mRNA levels were checked. Glycerol release into the medium was significantly increased by the cells treated with green tea extract. Glycerol release into the medium was significantly increased by the cells treated with green tea extract. Caffeine and theanine from green tea showed some level of lipolytic activity, and glycerol-3-phosphate dehydrogenase activity was remarkably decreased by EGCG. These results suggest that green tea has anti-obesity effect through inhibition of adipogenesis and stimulation of lipolysis. Catechins and theanine of green tea might be the factors responsible for the modulation of lipid metabolism and adipocyte differentiation.

• Applied Biological Chemistry
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• v.49 no.4
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• pp.339-342
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• 2006

Canna generalis was extracted with 80% aqueous MeOH, and the concentrated extract was partitioned with EtOAc, n-BuOH and $H_2O$, successively. from the EtOAc and n-BuOH fractions, four compounds were isolated through the repeated silica gel and ODS column chromatographies. From the results of physico-chemical data including NMR, MS and IR, the chemical structures of the compounds were determined as $\beta$-sitosterol(1), linoleic acid methyl ester(2),1-O-oleoyl-2-O-linoleoyl-3-O-$\beta$-D-galactopyranosyl-sn-glycerol(3), and daucosterol(4). They were the first to be isolated from Canna generalis.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.15 no.4
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• pp.303-316
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• 1993

The auricular cartilage grafts have been widely used in replacement of the temporomandibular joint disk. Cartilage grafts itself have a low metabolism and high survival rate after grafting. In processing the grafting materials, it was important to preserve the properties of chondrocyte proper. We used 15% glycerol and 10% DMSO (Dimethyl Sulfoxide) solutions for cartilage fixation before deep freezing. We have performed the allogenic auricular cartilage graft in the temporomandibular joint of 20 rabbits which 10 specimen was treated with 15% glycerol and the other 10 specimen was treated with 10% DMSO respectively and examined in 1, 2, 4, 6 and 8 weeks after operation histopathologically. The result were : 1. Inflammatory cell infiltration around the grafted material appeared more glycerol groups than DMSO groups at 1 week, but each group has no differences after 2 weeks. 2. Degenerative changes of grafted auricular chondrocytes were more deveolped in glycerol group than DMSO groups till 4 weeks, but there were no differences between two groups after 6 weeks. 3. Fibrous union between grafted fragment and mandibular condyle was prominent in DMSO group. 4. Vascular proliferation of the grafted auricualr cartilage was more developed in DMSO groups than glycerol group in early stage. 5. Amount of the additional growth of grafted auricular cartilage was more existed in DMSO groups than glycerol group. 6. General survival rate after grafting was more prominent in DMSO group. In summary, allogenic auricular cartilage grafts treated with 15% glycerol and 10% DMSO solution have supported to survivalbility as a cryopreservative agents, especially DMSO groups have little inflammatory cell infiltration in early stages and degenerative changes and additional growth are more prominent than glycerol groups.

• Journal of Microbiology and Biotechnology
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• v.26 no.6
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• pp.1077-1086
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• 2016

Glycerol dehydrogenases (GlyDHs) are essential for glycerol metabolism in vivo, catalyzing its reversible reduction to 1,3-dihydroxypropranone (DHA). The gldA gene encoding a putative GlyDH was cloned from Thermoanaerobacterium thermosaccharolyticum DSM 571 (TtGlyDH) and expressed in Escherichia coli. The presence of Mn²⁺ enhanced its enzymatic activity by 79.5%. Three highly conserved residues (Asp¹⁷¹, His²⁵⁴, and His²⁷¹) in TtGlyDH were associated with metal ion binding. Based on an investigation of glycerol oxidation and DHA reduction, TtGlyDH showed maximum activity towards glycerol at 60℃ and pH 8.0 and towards DHA at 60℃ and pH 6.0. DHA reduction was the dominant reaction, with a lower K_m(DHA) of 1.08 ± 0.13 mM and V_max of 0.0053 ± 0.0001 mM/s, compared with glycerol oxidation, with a K_m(glycerol) of 30.29 ± 3.42 mM and V_max of 0.042 ± 0.002 mM/s. TtGlyDH had an apparent activation energy of 312.94 kJ/mol. The recombinant TtGlyDH was thermostable, maintaining 65% of its activity after a 2-h incubation at 60℃. Molecular modeling and site-directed mutagenesis analyses demonstrated that TtGlyDH had an atypical dinucleotide binding motif (GGG motif) and a basic residue Arg⁴³, both related to dinucleotide binding.