• Journal of Microbiology and Biotechnology
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• v.30 no.8
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• pp.1261-1271
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• 2020

Cytochrome c_L (Cytc_L) is an essential protein in the process of methanol oxidation in methylotrophs. It receives an electron from the pyrroloquinoline quinone (PQQ) cofactor of methanol dehydrogenase (MDH) to produce formaldehyde. The direct electron transfer mechanism between Cytc_L and MDH remains unknown due to the lack of structural information. To help gain a better understanding of the mechanism, we determined the first crystal structure of heme c containing Cytc_L from the aquatic methylotrophic bacterium Methylophaga aminisulfidivorans MP^T at 2.13 Å resolution. The crystal structure of Ma-Cytc_L revealed its unique features compared to those of the terrestrial homologues. Apart from Fe in heme, three additional metal ion binding sites for Na⁺, Ca⁺, and Fe²⁺ were found, wherein the ions mostly formed coordination bonds with the amino acid residues on the loop (G93-Y111) that interacts with heme. Therefore, these ions seemed to enhance the stability of heme insertion by increasing the loop's steadiness. The basic N-terminal end, together with helix α4 and loop (G126 to Y136), contributed positive charge to the region. In contrast, the acidic C-terminal end provided a negatively charged surface, yielding several electrostatic contact points with partner proteins for electron transfer. These exceptional features of Ma-Cytc_L, along with the structural information of MDH, led us to hypothesize the need for an adapter protein bridging MDH to Cytc_L within appropriate proximity for electron transfer. With this knowledge in mind, the methanol oxidation complex reconstitution in vitro could be utilized to produce metabolic intermediates at the industry level.

• 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.

• Korean Journal of Microbiology
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• v.31 no.3
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• pp.179-183
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• 1993

A restricted facultatively methanol-oxidizing bacterium, Methylovorus sp. strain SS1, was isolate dfrom soil samples from Kuala Lumpur, Malaysia, through methanol-enrichment culture technique. The isolate was nonmotile Gram-negative rod and did not have complex internal membrane system. The colonies were small, pale-yellow, and raised convex with entire margin. The cell did not produce any spores and capsular materials. The cell was obligately aerobic and exhibited catalase, but no oxidase, activity. Plasmid, carotenoid pigment, and poly-.betha.-hydroxybutyric acid were not found. The guanine plus cytosine content of the DNA was 55%. The isolate was found to grow only on methanol methylamine, or glucose. Growth factors were not required. Cells growing on methanol was found to produce extracellular polysaccharides containing glucose, lactose, and fructose. Growth was optimal (t$_{d}$= 1.7) with 0.5%(v/v) methanol at 40.deg.C and pH 6.5. No Growth was observed at over 60.deg.C. Cell-free extracts of the methanol grown cells exhibited the phenazine methosulfate-linked methanol dehydrogenase activity Methanol was found to be assimilate dthrough the ribulose monophosphate pathway.y.

• YAKHAK HOEJI
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• v.34 no.6
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• pp.447-456
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• 1990

Various methods are available for the production of L-threonine. The microbial production of L-threonine has been achieved by breeding L-threonine analog-resistant auxotrophic mutants of various bacteria. The enzymatic production of L-threonine has been demonstrated by use of threonine metabolic enzymes such as threonine deaminase, threonine aldolase, or threonine dehydrogenase complex. Threonine synthesis from glycine and ethanol seems to be catalyzed by the enzymes Methanol dehydrogenase(MDH) and Serine hydroxymethyltransferase(SHMT), which was also found to catalyze the aldol condensation of glycine with acetaldehyde. The improved production of L-threonine has been achieved by amplifying the genes for the L-threonine biosynthetic enzymes using recombinant DNA techniques.

• Journal of Korean Society of Environmental Engineers
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• v.33 no.9
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• pp.662-669
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• 2011

This study was conducted to biologically convert methane into methanol. Methane contained in biogas was bio-catalytically oxidized by methane monooxygenase (MMO) of methanotrophs, while methanol conversion was observed by inhibiting methanol dehydrogenase (MDH) using MDH activity inhibitors such as phosphate, NaCl, $NH_4Cl$, and EDTA. The degree of methane oxidation by methanotrophs was the most highly accomplished as 0.56 mmol for the condition at $35^{\circ}C$ and pH 7 under 0.4 (v/v%) of biogas ($CH_4$ 50%, $CO_2$ 50%) / Air ratio. By the inhibition of 40 mM of phosphate, 50 mM of NaCl, 40 mM of $NH_4Cl$ and $150{\mu}m$ of EDTA, methane oxidation rate could achieve more than 80% regardless of type of inhibitors. In the meantime, addition of 40 mM of phosphate, 100 mM of NaCl, 40 mM of $NH_4Cl$ and $50{\mu}m$ of EDTA each led to generating the highest amount of methanol, i.e, 0.71, 0.60, 0.66, and 0.66 mmol when 1.3, 0.67, 0.74, and 1.3 mmol of methane was each concurrently consumed. At that time, methanol conversion rate was 54.7, 89.9, 89.6, and 47.8% respectively, and maximum methanol production rate was $7.4{\mu}mol/mg{\cdot}h$. From this, it was decided that the methanol production could be maximized as 89.9% when MDH activity was specifically inhibited into the typical level of 35% for the inhibitor of concern.