• Proceedings of the Korean Society of Food Hygiene and Safety Conference
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• 2002.05a
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• pp.215-215
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• 2002

Catalytic subunit of pyruvate dehydrogenase phosphatase (PDPc) has been suggested to have three major funational domains such as dihydrplipoamide adetyltransferase(E2)-binding domain, regulatory subunit of PDP(PDP)r-binding domain, and calcium-binding domain. In order to identify functional domains, recombinant catalytic subunit of pyruvate dehydrogenase phosphatase(rPDPc) was expressed in E. coli JM101 and purified to near homogeneity using the unique property of PDPc: PDPc binds to the inner lipoyl domain (L2) of E2 of ppyruvate dehydrogenase complex (PDC) in the presence of Ca+2, not under EGTA. PDPc was limited-proteolysed by typsin, chymotypsin, Arg-C, and elastase at pH 7.0 and 30C and N-terminal analysis of the fragments was done. Chymotrypsin, trypsin, and elastase made two major fragments: N-terminal large fragment, approx. 50kD and C-terminal small fragment, approx.10 kDa. Arg-C made three major fragments: N-terminal fragment, approx. 35kD, and central fragment, approx. 15 kD, and C-terminal fragment, approx. 10 kD. This study strongly suggest that PDPc consists of three major functional domains. However, further study should be necessary to identify the functional role.

• YAKHAK HOEJI
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• v.33 no.1
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• pp.39-45
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• 1989

Effects of dammarane glycosides of Panax ginseng on primary cultured chicken embryonic brain cells were studied by microscopic observation and determination of the activity of pyruvate dehydrogenase complex (PDHC). Brain cells were prepared from the brain of 10-day-old chicken embryo and cultured with either a standard medium consisted of 85% Dulbecco's Modified Eagle Medium (DMEM), 10% horse serum and 5% chicken embryonic extracts or a deficient medium consisted of 90% DMEM and 10% horse serum. It was observed that dammarane glycosides of Panax ginseng seemed to show the tendency to stimulate the neurite outgrowth of brain cells which were cultured with a deficient medium under microscopic observation. The activity of PDHC in brain cells cultured with a deficient medium was increased by dammarane glycosides of Panax ginseng.

• Korean Journal of Pharmacognosy
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• v.20 no.1
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• pp.32-36
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• 1989

Effects of Lycium chinensis, Epimedium koreanum and tuguaconitine which is isolated from Aconitum sibiricum on primary culture chicken embryonic brain cells were studied by microscopic observation and determined of the activity of pyruvate dehydrogenase complex(PDHC). Brain cells were prepared from the brain of 10-day-old chicken embryo and cultured with a medicine consisted of 90% Dulbecco's Modified Eagle Medium(DMEM) and 10% horse serum. It was observed that all substances studied seemed to show the tendency to stimulate the neurite outgrowth of brain cells which were cultured with a deficient medium under microscopic observation. The activity of PDHC in brain cells cultured with a deficient medium was increased by Lysium chinensis and Epimedium koreanum. However, tuguaconitine had not influence on the activity of PDHC.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1994.04a
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• pp.260-260
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• 1994

Mammalian pyruvate dehydrogenase complex(PDC) enzyme consists of multiple oopies of three major oligomeric enzymes-El, E2 E3. And protein X is one of the enzymatic constituents which is tightly bound to E2 subunit This complex enzyme is responsible for the oxidative decarboxylation of pyruvate producing of acetyl CoA which is a key intermediate for the entry of carbohydrates into the TCA cycle for its complete metabolic conversion to CO$_2$. And the overall activity of the complex enzyme is regulated via covalent nodification of El subunit by a El specific phosphatase ad kinase. Protein X has lipoyl moiety that undergoes reduction and acetylation during ezymatic reaction and has been known h be involved in the binding of E3 subunit to E2 core and in the regulatory activity of kinase. The purification of protein X has not been achieved majorly because of its tight binding to E2 subunit The E2-protein X subcomplex was obtained by the established methods and the detachment of protein X from E2 was accomplished in the 0.1M borate buffer containing 150mM NaCl. During the storage of the subcomplex in frozen state at -70$^{\circ}C$, the E2 subunit was precipitated and the dissociated protein X was obtained by cntrifegation into the supernatant The verification of protein X was accomplished by (1)the migration on SDS-PAGE, (2)acetylation by 〔2$\^$-l4/C〕 pyruvate, and (3)internal amino acid sequence analysis of tryptic digested enzyme.

• Archives of Pharmacal Research
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• v.14 no.4
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• pp.325-329
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• 1991

Effects of Lycii Fructus on primary cultured chicken embryonic brain cells were studied by microscopic observation, determination of the activity of pyruvate dehydrogenase complex (PDHC), and syntheses of protein, RNA and DNA. The brain cells were prepared from the brains or 10-day-old chicken embryos and cultured with a deficient medium. The activity of PDHC in the brain cells cultured with a deficient medium was increased to 1.8 times by the addition of $30\;{\mu}g/ml$ of the total methanol extract of Lycii Fructus. To seek the active fraction, total methanol extract was further fractionated by the polarity. The survival rate of neuronal cells was significantly increased by the addition of $100\;{\mu}g/ml$ of the buthanol or aqueous fraction. At this concentration, the significant increase of the syntheses of protein and RNA, but not of DNA, indicates that the fractions may act on the neuronal cells which are known to be non-dividing cells.

• The Korean Journal of Physiology and Pharmacology
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• v.9 no.4
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• pp.195-201
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• 2005

High extracellular glucose concentration was reported to suppress intracellular $Ca^{2+}$ clearing through altered sarcoplasmic reticulum (SR) function. In the present study, we attempted to elucidate the effects of pyruvate and fatty acid on SR function and reveal the mechanistic link with glucose-induced SR dysfunction. For this purpose, SR $Ca^{2+}$-uptake rate was measured in digitonin-permeabilized H9c2 cardiomyocytes cultured in various conditions. Exposure of these cells to 5 mM pyruvate for 2 days induced a significant suppression of SR $Ca^{2+}$-uptake, which was comparable to the effects of high glucose. These effects were accompanied with decreased glucose utilization. However, pyruvate could not further suppress SR $Ca^{2+}$-uptake in cells cultured in high glucose condition. Enhanced entry of pyruvate into mitochondria by dichloroacetate, an activator of pyruvate dehydrogenase complex, also induced suppression of SR $Ca^{2+}$-uptake, indicating that mitochondrial uptake of pyruvate is required in the SR dysfunction induced by pyruvate or glucose. On the other hand, augmentation of fatty acid supply by adding 0.2 to 0.8 mM oleic acid resulted in a dose-dependent suppression of SR $Ca^{2+}$-uptake. However, these effects were attenuated in high glucose-cultured cells, with no significant changes by oleic acid concentrations lower than 0.4 mM. These results demonstrate that (1) increased pyruvate oxidation is the key mechanism in the SR dysfunction observed in high glucose-cultured cardiomyocytes; (2) exogenous fatty acid also suppresses SR $Ca^{2+}$-uptake, presumably through a mechanism shared by glucose.

• Molecules and Cells
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• v.45 no.7
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• pp.454-464
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• 2022

DJ-1 is one of the causative genes of early-onset familial Parkinson's disease (PD). As a result, DJ-1 influences the pathogenesis of sporadic PD. DJ-1 has various physiological functions that converge to control the levels of intracellular reactive oxygen species (ROS). Based on genetic analyses that sought to investigate novel antioxidant DJ-1 downstream genes, pyruvate dehydrogenase (PDH) kinase (PDK) was demonstrated to increase survival rates and decrease dopaminergic (DA) neuron loss in DJ-1 mutant flies under oxidative stress. PDK phosphorylates and inhibits the PDH complex (PDC), subsequently downregulating glucose metabolism in the mitochondria, which is a major source of intracellular ROS. A loss-of-function mutation in PDK was not found to have a significant effect on fly development and reproduction, but severely ameliorated oxidative stress resistance. Thus, PDK plays a critical role in the protection against oxidative stress. Loss of PDH phosphatase (PDP), which dephosphorylates and activates PDH, was also shown to protect DJ-1 mutants from oxidative stress, ultimately supporting our findings. Further genetic analyses suggested that DJ-1 controls PDK expression through hypoxia-inducible factor 1 (HIF-1), a transcriptional regulator of the adaptive response to hypoxia and oxidative stress. Furthermore, CPI-613, an inhibitor of PDH, protected DJ-1 null flies from oxidative stress, suggesting that the genetic and pharmacological inhibition of PDH may be a novel treatment strategy for PD associated with DJ-1 dysfunction.

• KSBB Journal
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• v.15 no.5
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• pp.482-488
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• 2000

The Fab fraction of PDC-E2 specific human monoclonal antibody was produced using recombinant E. coli, and the effects of glucose and acetate were investigated to develop an optimal strategy for recombinant human antibody production. Higher glucose concentration in the culture media resulted inn higher cell growth and glucose consumption rate, which in turn resulted in an increased acetate production rate. When glucose was depleted, cells began to consume acetate as an energy source, and this consumption rate depended on the glucose concentration. When the residual glucose concentration was high, the accumulation of acetate was accelerated due to an increase in the acetate production rate and a decrease in the acetate consumption rate. Futhermore, it was found that a high accumulation of acetate, accompanied by a high glucose concentration, inhibited human antibody formation; the critical acetate concentration was $0.6g/\ell$. During production, a high glucose concentration enhanced cell growth, but inhibited antibody formation due to catabolic repression. Therefore, it is important to keep the concentration of both glucose and acetate as low as possible to increase antibody production after induction. Accordingly, it is important to accurately control the concentration of glucose and acetate in the culture media to obtain high cell densities and high productivity levels of recombinant human antibody.

• 한국체육학회지인문사회과학편
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• v.55 no.3
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• pp.649-658
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• 2016

This study aimed to analyze PDH(Pyruvate dehydrogenase) and protein expression of PDK4(Pyruvate dehydrogenase kinase 4), PDP1(PDH phosphatase 1), enzymes that are involved in the activation of PDH, in skeletal muscle and to investigate the concentration of thiamine administration in liver and muscle following 4 weeks of endurance training. Methods : 6 weeks old male ICR mice were divided into two groups: sedentary group (CON, n=10; TH, n=10), and exercise group (EX, n=10, THEX, n=10). Thiamine(thiamine tetrahydrofurfuryl disulfide: TTFD) TTFD was orally administrated into TH and THEX groups in 50mg/kg body weight for 4 weeks. Treadmill training was performed in EX and THEX groups at about 70% of VO2max for 5 times a week for 4 weeks. Results : In this study, the concentration of glycogen was significantly increased following 4 weeks of endurance training, but a significant difference was not found following thiamine administration. Similarly, there was a significant effect of the training on PDH and the expression of PDK4 and PDP1 as PDH was increased by about 40% along with the increase in PDK4 and PDP1. However, there was no significant difference found between the groups following thiamine administration. Discussion : This result shows that there was no synergistic effect of thiamine administration, potentially due to adaptation of skeletal muscle from a long-term endurance training. Therefore, it will be necessary to consider the intake timing of thiamine and to analyze proteins that are related to PDH following the administration of complex carbohydrates.

• KSBB Journal
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• v.15 no.5
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• pp.489-496
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• 2000

Several culture systems including batch, two-stage CSTR, semi-fed batch, and two-stage cyclic fed-batch were investigated for the efficient production of the Fab fraction of PDC-E2 specific human monoclonal antibody using high cell density recombinant E. coli. A two-phase batch system and a two-stage continuous system were examined to overcome plasmid instability problems, by separating the growth and the production stages. The cell density and productivity of the two-stage continuous culture was better than that of the two-phase batch fermentation. In the two-stage continuous culture system with DO-stat, the cell growth and the productivity were superior to those of the system without the DO control. Also, almost total plasmid stability was maintained in the two-stage continuous culture system. Modified M9 medium was selected as an optimum feeding medium for the fed-batch process, and the optimum C/N ratio determined to be 2:3. The optimum feeding rate was $0.6g/\ell/hr$ for a constant feeding strategy in semi-fed batch system. When the feeding medium was fed by pulsing, it was observed that more frequent pulsing resulted in improved cell growth. The linear feeding method was the most efficient of the various feeding methods tested. Finally, high cell density culture using a two-stage cyclic fed batch system with pH-stat was tried because the linear feeding method showed limitations in terms of obtaining high cell densities, and a cell density of $54 g/\ell$ was achieved. It was concluded that the two-stage cyclic fed batch system was the most efficient system for high cell density culture of the systems tested. However, productivity improvements were lower than expected due to the extremely high accumulations of acetate, although the low levels of residual glucose were maintained.