• BMB Reports
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• v.44 no.9
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• pp.590-594
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• 2011

The production of antileukemic enzyme methionine ${\gamma}$-lyase (MGL) in distinctly related bacteria, Citrobacter freundii and in their recombinants expressing the Vitresocilla hemoglobin (VHb) has been studied. This study concerns the potential of Citrobacter freundii expressing the Vitreoscilla hemoglobin gene (vgb) for the methionine ${\gamma}$- liyase production. Methionine ${\gamma}$- liyase production by Citrobacter freundii and its $vgb^-$ and $vgb^+$ bearing recombinant strain was studied in shake-flasks under 200 rpm agitation, culture medium and $30^{\circ}C$ in a time-course manner. The $vgb^+$ and especially the carbon type had a dramatic effect on methionine ${\gamma}$- liyase production. The $vgb^+$ strain of C. freundii had about 2-fold and 3.1-fold higher levels of MGL than the host and $vgb^-$ strain, respectively.

• Journal of Microbiology and Biotechnology
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• v.14 no.2
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• pp.373-378
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• 2004

The regulatory mechanism of methionine biosynthesis in Corynebacterium glutamicum was analyzed at the protein arid gene expression level. O-Acetylhomoserine sulfhydraylase (encoded by metY) was inhibited by 10 mM methionine to a residual activity of 10% level, whereas no such inhibition was found with cystathionine $\gamma$-synthase (encoded by metB) and cystathionine $\beta$-lyase (encoded by metC). The enzymatic activity of homoserine acetyltransferase (encoded by metX) was repressed to a residual activity of 25% level by 10 mM methionine which was added to the growth medium. Cystathionine $\gamma$-synthase and cystathionine $\beta$-lyase were also repressed by 10 mM methionine, but only to a residual activity of 50-70% level. O-Acetylhomoserine sulfhydrylase was very sensitive to repression by 10 mM methionine, showing residual activity of 13%. In addition, homoserine acetyltransferase was also repressed by 10 mM cysteine to 50% of its original activity. No repression of the enzymes by S-adenosyl methionine was observed. The pattern of repression by methionine indicated that the metB and aecD genes might be regulated by a common mechanism, while the metA and metY genes are differently regulated.

• Microbiology and Biotechnology Letters
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• v.42 no.1
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• pp.11-17
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• 2014

Selenium is an essential trace element for mammals, but it is very toxic. Therefore, the control of selenium concentrations should be precisely and effectively monitored. Selenium is naturally obtained through foods and seleno-L-methionine (LSeMet) is a major form of selenium. It has been reported that L-SeMet is only converted into Se-adenosyl-L-SeMet. However, a recent study suggested that L-SeMet was directly metabolized into methylselenol ($CH_3SeH$) in mouse liver extract by the reaction of cystathionine ${\gamma}$-lyase (CGL). The canonical reaction of CGL was known to catalyze the cleavage of L-cystathionine to L-cysteine, ${\alpha}$-ketobutyrate and $NH_3$. In the present study, we found that L-SeMet could be directly converted to $CH_3SeH$ using purified homogenous human CGL instead of mouse liver cytosol. Authentic $CH_3SeH$ was prepared by reduction of dimethyldiselenide with sodium tetrahydroborate. The gaseous product of the enzymatic reaction with L-SeMet was analyzed by GC/MS spectrometry. The GC/MS data was identical to that of authentic dinitrophenyl selenoether. We also analyzed the kinetic parameters for the formation of $CH_3SeH$ from L-SeMet by human and mouse CGL. These results suggest that human CGL is a critical enzyme which is responsible for L-SeMet metabolism.

• Journal of Microbiology and Biotechnology
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• v.23 no.4
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• pp.499-510
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• 2013

Among 25 isolates, Aspergillus fumigatus ASH (JX006238) was identified as a potent producer of homocysteine ${\gamma}$-lyase. The nutritional requirements to maximize the enzyme yield were optimized under submerged (SF) and solid-state fermentation (SSF) conditions, resulting in a 5.2- and 2.3-fold increase, respectively, after the last purification step. The enzyme exhibited a single homogenous band of 50 kDa on SDS-PAGE, along with an optimum pH of 7.8 and pH stability range of 6.5 to 7.8. It also showed a pI of 5.0, as detected by pH precipitation with no glycosyl residues. The highest enzyme activity was obtained at $37-40^{\circ}C$, with a $T_m$ value of $70.1^{\circ}C$. The enzyme showed clear catalytic and thermal stability below $40^{\circ}C$, with $T_{1/2}$ values of 18.1, 9.9, 5.9, 3.3, and 1.9 h at $30^{\circ}C$, $35^{\circ}C$, $40^{\circ}C$, $50^{\circ}C$, and $60^{\circ}C$, respectively. Additionally, the enzyme $K_r$ values were 0.002, 0.054, 0.097, 0.184, and 0.341 $S^{-1}$ at $30^{\circ}C$, $35^{\circ}C$, $40^{\circ}C$, $50^{\circ}C$, and $60^{\circ}C$, respectively. The enzyme displayed a strong affinity to homocysteine, followed by methionine and cysteine when compared with non-S amino acids, confirming its potency against homocysteinuria-related diseases, and as an anti-cardiovascular agent and a specific biosensor for homocysteinuria. The enzyme showed its maximum affinity for homocysteine ($K_m$ 2.46 mM, $K_{cat}\;1.39{\times}10^{-3}\;s^{-1}$), methionine ($K_m$ 4.1 mM, $K_{cat}\;0.97{\times}10^{-3}\;s^{-1}$), and cysteine ($K_m$ 4.9 m M, $K_{cat}\;0.77{\times}10^{-3}\;s^{-1}$). The enzyme was also strongly inhibited by hydroxylamine and DDT, confirming its pyridoxal 5'-phosphate (PLP) identity, yet not inhibited by EDTA. In vivo, using Swiss Albino mice, the enzyme showed no detectable negative effects on platelet aggregation, the RBC number, aspartate aminotransferase, alanine aminotransferase, or creatinine titer when compared with negative controls.

• International Journal of Oral Biology
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• v.44 no.4
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• pp.152-159
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• 2019

The oral care probiotic strain Weissella cibaria CMU (oraCMU) inhibits volatile sulphur compounds associated with halitosis, presumably by inhibiting the growth of associated oral pathogens. In the present study, we investigated whether oraCMU inhibits the production of these compounds by suppressing the expression of mgl. This gene encodes L-methionine-α-deamino-γ-mercaptomethane-lyase (METase) and is involved in the production of methyl mercaptan (CH₃SH) by Porphyromonas gingivalis. Therefore, we specifically investigated the effects of oraCMU on the growth, CH₃SH production, METase activity, and mgl expression of P. gingivalis. The minimum inhibitory concentrations of cell-free supernatant and secreted proteins from oraCMU were 125 mg/mL and 800 ㎍/mL, respectively. At sub-minimum inhibitory concentration levels, these metabolites inhibited CH₃SH production, but they also reduced P. gingivalis viability. Only heat-killed oraCMU decreased CH₃SH production without affecting P. gingivalis viability. Heat-killed oraCMU also inhibited METase activity toward L-methionine and mgl mRNA expression (p < 0.05). In summary, we demonstrated the inhibition of volatile sulphur compounds via the antimicrobial action of oraCMU and, for the first time, the inhibition of such compounds by heat-killed oraCMU, which occurred at the molecular level.

• Food Science and Biotechnology
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• v.14 no.5
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• pp.621-627
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• 2005

Halitosis is mainly caused by the presence of volatile sulfur-containing compounds (VSC's) produced by proteolytic periodontopathic bacteria in the oral cavity. Various mouth-rinses have been offered on the market as solutions to reduce halitosis. The aim of this study was to find a potent substance for the prevention of halitosis. The halitosis prevention substance (HPS) from cumin seed powder was purified by solvent extraction, silica gel column chromatography and preparative TLC to yield an oil phase (0.98%). Instrumental analysis such as FT-IR, $^1H$-NMR and $^{13}C$-NMR showed that HPS contained an -OH group, -HC=CH-, -COO-, and long chain acyl group. HPS was therefore determined to be 2-hydroxyethyl-${\beta}$-undecenate. HPS inhibited the growth of Fusobacterium nucleatum and Porphyromonas gingivalis, by 72.44% and 64.37% at $1{\times}10^{-2}\;M$, and by 99.85% and 91.62% at $5\;{\times}\;10^{-2}\;M$, respectively. It also inhibited the activity of L-methionine-${\alpha}$-deamino-${\gamma}$-mercaptomethane-lyase (METase), which was produced by oral microbes. Furthermore, the VSC production by oral microbes in the human mouth air decreased with increasing HPS concentration. These results suggested that HPS from cumin seed is an efficient halitosis prevention agent.