• KSBB Journal
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• v.26 no.3
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• pp.189-192
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• 2011

Much interest has been recently focused on the production of large quantities of hydrogen, due to its potential importance in our economy and needs in the petroleum and chemical industries. Formate dehydrogenase H (FDH-H) from Escherichia coli containing selenocysteine that oxidizes formate to carbon dioxide with the release of hydrogen is a component of the anaerobic formate hydrogen-lyase complex of E. coli. To make full use of FDH-H, we need effective expression condition. In this approach, we investigated the effect of pH on FDH-H stability and observed the effect of selenite and formate concentration on the activity of FDH-H. Additionally, coexpression of selenocysteine insertion genes were tried to improve the expression of FDH-H. The highest level of FDH-H expression was achieved by coexpression of selenocysteine insertion genes (pSUABC) as well as by the addition of $10\;{\mu}M$ selenite and 10 mM formate. At this optimized condition, a 2.6 fold elevation of expression of FDH-H was achieved.

• Proceeding of EDISON Challenge
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• 2013.04a
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• pp.67-75
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• 2013

최근 이상기후의 원인으로 손꼽히는 물질의 중심에는 이산화탄소가 있으며 이를 제거하기 위한 여러 연구가 진행되고 있다. 최근에는 전극이 있는 수조에 미생물을 넣고 이산화탄소를 화학적 에너지로 사용할 수 있도록 알코올로 변환시켜주는 시스템이 발표되었다. 이에 따라 본 연구진에서는 이러한 전극 시스템에서 이용될 수 있는 효소를 찾고 효소촉매화 반응의 메커니즘을 자세히 연구하고자 하였다. 본 연구에서 사용된 효소인 Formate dehydrogenase (FDH)는 formate를 조효소인 nicotinamide adenine dinucleotide ($NAD^+$)를 사용하여 이산화탄소로 산화시키는 반응을 촉진시키는 효소이다. 본 연구에서는 이러한 FDH의 산화반응의 역반응을 이용하여 이산화탄소를 효과적으로 분해하는 메커니즘을 연구하기에 앞서 wild type의 반응 메커니즘에 대해 깊이 연구하고자 B3LYP 방법의 양자계산을 하여 반응의 transition state와 potential energy를 조사하였다.

• Journal of the Korean Wood Science and Technology
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• v.50 no.3
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• pp.159-166
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• 2022

CO₂ emissions are the primary reason for global warming; hence, biological and chemical technologies for converting CO₂ into useful compounds are being actively studied. Biological methods using enzymes can convert CO₂ under mild conditions. Formate dehydrogenase (FDH) is a representative CO₂ conversion enzyme. Its function was revealed after isolation from bacteria, yeast, and plants. In this study, we evaluated the CO₂ conversion potential of FDH isolated from wood-rotting fungi. After isolating the FDH gene (TvFDH) from Trametes versicolor, we cloned the full-length FDH from T. versicolor and expressed it in a cell-free expression system. The gene encoding TvFDH was identified as 1,200 bp open reading frame (ORF) and the expected molecular weight of the protein was approximately 42 kDa. Overexpression of the recombinant crude protein including TvFDH was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Enzyme activities and metabolite analyses confirmed the efficiency of TvFDH for CO₂ reduction.

• Journal of Korean Society of Forest Science
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• v.102 no.3
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• pp.331-337
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• 2013

Formate dehydrogenase (FDH), catalyzing the oxidation of the formate ion to carbon dioxide, is known as the stress protein in response to drought, low temperature and pathogen infection. To study the functions of FDH in poplar (Populus alba ${\times}$ P. glandulosa), we isolated a FDH cDNA (PagFDH1) and examined its expressional characteristics. The PagFDH1 is 1,499 base pairs long and encodes a putative 388 amino acid protein with an expected molecular mass of 42.5 kDa. The PagFDH1 protein has N-terminal mitochondria signal peptide and $NAD^+$ binding domain. Southern blot analysis indicated that a single copy of the PagFDH1 is present in the poplar genome. PagFDH1 is expressed highly in the suspension cells (especially in the lag and early exponential phases) and moderately in roots, flowers and leaves. ABA-mediated enhanced expression of PagFDH1 in response to drought and salt stress treatments indicates that the gene product could play an important role in the development of stress resistant trees.

• Applied Chemistry for Engineering
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• v.20 no.4
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• pp.449-452
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• 2009

The absence of Fe, Se, and Mo in a minimal medium prevented the production of hydrogen from the anaerobic culture of Escherichia coli MC4100. Fe, Se, and Mo are known to be cofactors of formate dehydrogenase ($FDH_{II}$) of both E. coli and Enterobacter aerogenes. Hence when these trace elements are absent in the minimal medium, hydrogen production through formate dehydrogenation would be inhibited not only in E. coli but also in E. aerogenes. Hydrogen production by E. aerogenes 413 was delayed when lacking these trace elements. Therefore, it is believed that hydrogen production of E. aerogenes is initiated not by the reoxidation of nicotinamide adenine dinucleotide (NADH) but by formate decarboxylation.

• Microbiology and Biotechnology Letters
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• v.25 no.4
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• pp.403-407
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• 1997

Medium components for maximum activity of NAD$^{+}$-dependent formate dehydrogenase (EC 1.2.1.2; FDH) were optimized with a methanol-assimilating yeast Hansenula sp. MS-364, preserved by our laboratory. The maximum activity of the enzyme was obtained when the strain was cultivated at 30$circ$C for 24 hours in a medium containing methanol 3%(v/v), yeast extract 0.8%(w/v), K$_{2}$HPO$_{4}$, 0.1%(w/v), KH$_{2}$PO$_{4}$ 0.1%(W/V), MgSO$_{4}$, 7H$_{2}$O 0.05%(w/v), and the pH of the culture broth was adjusted at 5.0.

• Journal of Microbiology and Biotechnology
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• v.13 no.4
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• pp.501-508
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• 2003

Artificial coupling of one enzyme with another can provide an efficient means for the production of industrially important chemicals. Xylose reductase has been recently discovered to be useful in the reductive production of xylitol. However, a limitation of its in vitro or in vivo use is the regeneration of the cofactor NAD(P)H in the enzyme activity. In the present study, an efficient process for the production of xylitol from D-xylose was established by coupling two enzymes. A NADH-dependent xylose reductase (XR) from Pichia stipitis catalyzed the reduction of xylose with a stoichiometric consumption of NADH, and the resulting cofactor $NAD^+$ was continuously re-reduced by formate dehydrogenase (FDH) for regeneration. Using simple kinetic analyses as tools for process optimization, suitable conditions for the performance and yield of the coupled reaction were established. The optimal reaction temperature and pH were determined to be about $30^{\circ}C$ and 7.0, respectively. Formate, as a substrate of FDH, affected the yield and cofactor regeneration, and was, therefore, adjusted to a concentration of 20 mM. When the total activity of FDH was about 1.8-fold higher than that of XR, the performance was better than that by any other activity ratios. As expected, there were no distinct differences in the conversion yields of reactions, when supplied with the oxidized form $NAD^+$ instead of the reduced form NADH, as a starting cofactor for regeneration. Under these conditions, a complete conversion (>99%) could be readily obtained from a small-scale batch reaction.

• Korean Chemical Engineering Research
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• v.54 no.5
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• pp.719-721
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• 2016

Formate has been considered as an environmentally sustainable feedstock that can be used to accelerate the production of valuable chemicals. This study presents brief results of the formatotrophic production of Poly-${\beta}$-hydroxybutyric acid (PHB) by Methylobacterium sp. To evaluate the production of PHB, five species of Methylobacteria were tested using formate as the sole carbon and energy source. Methylobacterium chloromethanicum CM4 exhibited the highest productivity of PHB, which showed 1.72 g/L PHB production, 32.4% PHB content, and 0.027 g-PHB/g-formate PHB yield. These results could be used for the formatotrophic production of PHB with the concurrent reduction of $CO_2$ to formate.

• Microbiology and Biotechnology Letters
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• v.34 no.3
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• pp.216-220
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• 2006

Thermophillic fungus Thermoascus aurantiacus showed excellent growth and produced high amount of alcohol oxidase and catalase in a pectin medium. Besides, the strain produced enzymes which related with pectin or alcohol decomposition. We detected extracellular pectin esterase (EC 3.1.1.11) activity and, both intracellular and extracellular pectinase (EC 4.2.2.10) activity, as pectinolytic enzymes produced by T. aurantiacus. The production of methanol decomposition enzymes, such as alcohol oxidase (AOD, EC 1.1.3.13), alcohol dehydrogenase (ADH, EC 1.1.1.1), formaldehyde dehydrogenase (FADH, EC 1.2.1.1) and formate dehydrogenase (FDH, EC 1.2.1.2) follows by pectin esterase reaction which is converted to methanol. We concluded that T. aurantiacus has pectinolytic and alcohol - oxidative enzymological mechanism which produced carbon dioxide as a final material, started from pectin.

• YAKHAK HOEJI
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• v.47 no.3
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• pp.184-189
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• 2003

Peptide deformylase (PDF) is essential and unique to bacteria, thus making it an attractive target for the discovery of novel antibacterial drugs. PDF deformylates the N-formylmethionine of newly synthesized polypeptides in prokaryotes. In this study, a pdf gene from Staphylococcus aureus 6538p was cloned in pET-14b vector and PDF protein was over-produced in Escherichia coli BL21 (DE3). NH$_2$-terminal His-tagged PDF protein was purified by nickel-nitrilotriacetic acid (Ni-NTA) metal-affinity chromatography. Enzymatic activity of purified 6xHis-tagged PDF was tested on the substrate (formyl-Methionine-Alanine-Serine) by formate dehydrogenase-coupled spectrometric assay of peptide deformylase. For the discovery of new PDF inhibitors from chemical libraries and culture broths of soil bacteria, a target-oriented screening system using a 96-well plate was developed. About 3,000 commercial chemical libraries were tested in this screening system, and 2 chemicals (0.07％) among them showed an inhibitory activity against PDF enzyme. This result showed that a new screening system can be used for the discovery of new PDF inhibitors.