• 한국생물공학회:학술대회논문집
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• 2003.04a
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• pp.587-587
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• 2003

• Proceedings of the Korean Society of Plant Biotechnology Conference
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• 2005.04a
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• pp.174-174
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• 2005

• 대한임상약리학회:학술대회논문집
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• 1999.12a
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• pp.30-30
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• 1999

• Proceedings of the Korean Society of Applied Entomolohy Conference
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• 2005.05a
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• pp.186-186
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• 2005

• Journal of Microbiology and Biotechnology
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• v.30 no.1
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• pp.146-154
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• 2020

With the advantages of biocatalytic method, enzymes have been excavated for the synthesis of chiral amino acids by the reductive amination of ketones, offering a promising way of producing pharmaceutical intermediates. In this work, a robust phenylalanine dehydrogenase (PheDH) with wide substrate spectrum and high catalytic efficiency was constructed through rational design and active-site-targeted, site-specific mutagenesis by using the parent enzyme from Bacillus halodurans. Active sites with bonding substrate and amino acid residues surrounding the substrate binding pocket, 49L-50G-51G, 74M,77K, 122G-123T-124D-125M, 275N, 305L and 308V of the PheDH, were identified. Noticeably, the new mutant PheDH (E113D-N276L) showed approximately 6.06-fold increment of k_cat/Km in the oxidative deamination and more than 1.58-fold in the reductive amination compared to that of the wide type. Meanwhile, the PheDHs exhibit high capacity of accepting benzylic and aliphatic ketone substrates. The broad specificity, high catalytic efficiency and selectivity, along with excellent thermal stability, render these broad-spectrum enzymes ideal targets for further development with potential diagnostic reagent and pharmaceutical compounds applications.

• KSBB Journal
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• v.19 no.5
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• pp.394-398
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• 2004

A derivatization method in mass spectrometry for small molecular analysis was developed to solve the problems of volatility of many analytes, difficult ionization of analytes, and undiscriminating isobaric analytes. This derivatization method, oximation of $\alpha$-ketoacid, in the transaminase reaction leads to change of mass difference between amine reactants and ketoacid products. In addition, regardless of the kinds of ketoacid, the linear relationship between the peak intensity and its concentration (from 1 mM to 10 mM) shows that quantitative analysis of the conversion of the reaction can be executed by the analysis of peak intensity of the corresponding oximated ketoacids. Furthermore, this method can be used for identifying transaminase as well as determining its substrate specificity

• BMB Reports
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• v.28 no.5
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• pp.451-457
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• 1995

Four carboxypeptidases, CP1, CP2, CP3, and CP4 were isolated from the cotyledons of germinating seedlings of Canavalia lineata by sequential chromatography on the following four columns: 1) CM-cellulose, 2) Sephacryl 5-300, 3) Procion red dye, and 4) Sephacryl S-200. A number of properties of the enzymes, such as substrate specificity, molecular weight, optimum pH, thermal stability, have been determined. Enzyme activities were measured using the Cbz(carbobenzoxy)-dipeptides containing phenylalanine at the penultimate position. The $K_m$ values of four carboxypeptidases for Cbz-Phe-Ala were 0.50, 0.65, 1.30, and 1.35 mM, respectively. The inhibition studies indicated that the four carboxypeptidases were all serine type. Each of the carboxypeptidases with molecular weights of 145, 114, 105, and 104 kDa, respectively, had the optimum enzyme activity at pH 5.0~6.0. And they were sensitive to high temperature.

• Journal of Microbiology and Biotechnology
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• v.12 no.2
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• pp.242-248
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• 2002

Previously, it was reported that the nonhomologous C-terminal regions of the D-hydantoinases are nonessential for catalysis, but affect the oligomeric structure of the enzyme [3]. In an effort to further confirm the above observation, the C-terminal region-inserted enzyme was constructed by attaching a peptide (22 residues) at the C-terminal of the D-hydantoinase from Bacillus thermocatenulatus GH2, and its structural and biochemical properties were compared with both the wild-type and C-terminal region-truncated enzymes. As a result, native tetrameric D-hydantoinase was dimerized as the truncated enzyme, and the inserted mutant with a new sequence was expressed as a catalytically active form in E. coli. Expression level of the inserted and truncated enzymes were found to be significantly increased compared to the level of the wild-type enzyme, and this appears to be due to the reduced toxic effect of the mutant enzymes on host cells. Dimerized enzymes exhibited increased thermo- and pH stabilities considerably when compared with the corresponding wild-type enzyme. Comparison of the substrate specificity between the mutant and wild-type enzymes suggests that the substrate specificity of the D-hydantoinase is closely linked with the oligomeric structure.

• Journal of the Korean Chemical Society
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• v.44 no.5
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• pp.448-452
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• 2000

Phospholipase D(PLD) is the enzyme catalyzing the hydrolysis of the terminal phosphcester bond of phospholipid head group to produce phosphatidic acid and the corresponding base. The effect of spermine on the PLD activity of rat brain mitochondrial preparation was investigated. Spermine, in the presence of oleic acid, activates the rat brain mitochondrial PLD, whose effect was further enhanced by the presence of divalent cation, $Ca^{2+}$, $Mg^{2+}$, and $Ba^{2+}$. Among the various monoamines tested, only histamine at the high concentration was effective in activation the PLD. Polylysine increased the PLD activity, particularly, the longer chain of the molecule activated the PLD more effectively. There was no significant difference in the substrate specificity for the PLD activity between phosphatidylcholine(PC) and phoshpatidylethanolamine (PE). This substrate specifitiy is different from the PE specificity reported for the intestinal mitochondrial PLD.

• Korean Journal of Microbiology
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• v.26 no.3
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• pp.207-214
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• 1988

A cytosine deaminase from the cell-free extract of an isolate was examined after ethyl alcohol reactionation. The enzyme catalyzed the conversion of 5-fluorocytosine to 5-fluorouracil by the possession of specificity to the substrate. The optimum temperature and storage time on the stability of the enzyme were at below $50^{\circ}C$ and near 2 days in tris-HCl buffer. The maximum activity was also presented ar 9.0 in pH and $45^{\circ}C$ in temperature. The pHs and temperatures for the enzyme activity ranged from 8.5-9.5 and from 40-$50^{\circ}C$, respectively. the presence of $Ag^{+}, Hg^{2+}, Zn^{2+}$ in the reaction mixture resulted in the marked inhibition in the activity, but 1mM of $Fe^{3+}, K^{+}$, or $Na^{+}$ increased the enzyme activity. The enzyme preparation was vot affected by inhibitors used except N-ethylmaleimide of 1 and 10mM, and considerably activated by 1mM of pyrophosphate and 10mM of phosphate.