• Journal of Microbiology and Biotechnology
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• v.2 no.3
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• pp.226-229
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• 1992

An ethanol tolerant mutant was selected by successive transfers of Clostridium thermohydrosulfuricum ATCC 33223 into the media with progressively higher ethanol concentrations. The growth kinetics of the mutant were characterized under various growth conditions. Physiological differences such as enhanced growth, tolerance to various solvents, alteration of the optimum temperature and the ratio of end products during fermentation were noticed in the mutant.

• Proceedings of the Korean Society of Crop Science Conference
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• 2017.06a
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• pp.174-174
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• 2017

To cope with phosphate (Pi) deficient stress, plants modulate various physiological and developmental processes, such as gene expression, Pi uptake and translocation, and root architecture changes. Here, we report the identification and characterization of novel activation-tagged mutant involved in Pi starvation signaling in Arabidopsis. The hpd (${\underline{h}ypersensitive}$ to ${\underline{P}i}$ $ {\underline{d}eficiency}$) mutant exhibits enhanced phosphate uptake and altered root architectural change under Pi starvation compared to wild type. Expression analysis of auxin-responsive DR5::GUS reporter gene in hpd mutant indicated that auxin translocation in roots under Pi starvation are suppressed in hpd mutant plants. Impaired auxin translocation in roots of hpd mutant was attributable to abnormal root architecture changes in Pi starvation conditions. Our results indicated that abnormal auxin translocation in hpd mutant might be due to mis-regulation of auxin efflux carrier proteins, PIN-FORMED (PIN) 1, and 2 under Pi starvation conditions. Not only expression levels but also expression domains of PIN proteins were altered in hpd mutant in response to Pi starvation. Molecular genetic analysis of hpd mutant revealed that the mutant phenotype is caused by the lesion in ENHANCED SILENCING PHENOTYPE4 (ESP4) gene whose function is proposed in mRNA 3'-end processing. The results suggest that mRNA processing plays crucial roles in Pi homeostasis as well as developmental reprograming in response to Pi deprivation in Arabidopsis.

• Journal of Microbiology and Biotechnology
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• v.26 no.12
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• pp.2066-2075
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• 2016

In this study, an enhanced lipid-producing mutant strain of the microalga Chlamydomonas reinhardtii was developed by gamma irradiation. To induce the mutation, C. reinhardtii was gamma irradiated at a dose of 400 Gy. After irradiation, the surviving cells were stained with Nile red. The mutant (Cr-4013) accumulating 20% more lipid than the wild type was selected. Thin-layer chromatography revealed the triglyceride and free fatty acid contents to be markedly increased in Cr-4013. The major fatty acids identified were palmitic acid, oleic acid, linoleic acid, and linolenic acid. Random amplified polymeric DNA analysis showed partial genetic modifications in Cr-4013. To ascertain the changes of protein expression in the mutant strain, two-dimensional electrophoresis was conducted. These results showed that gamma radiation could be used for the development of efficient microalgal strains for lipid production.

• Journal of Microbiology and Biotechnology
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• v.17 no.5
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• pp.721-727
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• 2007

Stability-enhanced mutants, H44, 11-94, 5A2-84, and F8, of L-threonine aldolase(L-TA) from Streptomyces coelicolor A3(2)(SCO1085) were isolated by an error-prone PCR followed by a high-throughput screening. Each of these mutant, had a single amino acid substitution: H177Y in the H44 mutant, A169T in the 11-94 mutant, D104N in the 5A2-84 mutant and F18I in the F8 mutant. The residual L-TA activity of the wild-type L-TA after a heat treatment for 20 min at $60^{\circ}C$ was only 10.6%. However, those in the stability-enhanced mutants were 85.7% for the H44 mutant, 58.6% for the F8 mutant, 62.1% for the 5A2-84 mutant, and 67.6% for the 11-94 mutant. Although the half-life of the wild-type L-TA at $63^{\circ}C$ was 1.3 min, those of the mutant L-TAs were longer: 14.6 min for the H44 mutant, 3.7 min for the 11-94 mutant, 5.8 min for the 5A2-84 mutant, and 5.0 min for the F8 mutant. The specific activity did not change in most of the mutants, but it was decreased by 45% in the case of mutant F8. When the aldol condensation of glycine and 3,4-dihydroxybenzaldehyde was studied by using whole cells of Escherichia coli containing the wild-type L-TA gene, L-threo-3,4-dihydroxyphenylserine(L-threo-DOPS) was successfully synthesized with a yield of 2.0 mg/ml after 20 repeated batch reactions for 100 h. However, the L-threo-DOPS synthesizing activity of the enzyme decreased with increased cycles of the batch reactions. Compared with the wild-type L-TA, H44 L-TA kept its L-threo-DOPS synthesizing activity almost constant during the 20 repeated batch reactions for 100 h, yielding 4.0 mg/ml of L-threo-DOPS. This result showed that H44 L-TA is more effective than the wild-type L-TA for the mass production of L-threo-DOPS.

• Journal of Pharmaceutical Investigation
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• v.33 no.4
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• pp.267-272
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• 2003

Herpes simplex virus (HSV) thymidine kinase (TK) has been widely used for suicidal gene therapy in combination with nucleoside analogs such as ganciclovir (GCV). The use of HSV-TK is limited due to the side effect of GCV at high concentrations. Previous studies showed that stable transfectants of mutant HSV-TK with enhanced affinity to GCV were killed at lower GCV concentrations. In this study, we tested whether mutant HSV-TK can provide enhanced suicidal effect when transiently transfected with Epstein-Barr virus (EBV)-based plasmid. EBV-based plasmid which contains OriP and EBNA-1 sequence is well known for a stable episomal maintenance in human cells. Optimal transfection condition was assessed for SNU-638 gastric cancer cell line using polyetylnimine (PEI). Maximum transfection efficiency was achieved when DNA:PEI was 1:3 (w/v). Cytotoxicities of mutant and wild type HSV-TK were compared before and after partially selecting transfected cells. The cells were sensitive to $100\;{\mu}g/ml$ hygromycin. Following GCV treatment, more cells were killed after hygromycin selection than before selection. The mutant HSV-TK showed enhanced cytotoxicity compared with the wild type HSV-TK. Our results suggest that the EBV-based plasmid encoding mutant HSV-TK may be useful to treat the diseases caused by uncontrolled cell proliferation such as cancer and rheumatoid arthritis.

• BMB Reports
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• v.44 no.2
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• pp.102-106
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• 2011

Thermodynamic stability and refolding kinetics of firefly luciferase and three representative mutants with depletion of negative charge on a flexible loop via substitution of Glu by Arg (ER mutant) or Lys (EK mutant) as well as insertion of another Arg in ER mutants (ERR mutant) was investigated. According to thermodynamic studies, structural stability of ERR and ER mutants are enhanced compared to WT protein, whereas, these mutants become prone to aggregation at higher temperatures. Accordingly, it was concluded that enhanced structural stability of mutants depends on more compactness of folded state, whereas aggregation at higher temperatures in mutants is due to weakening of intermolecular repulsive electrostatic interactions and increase of intermolecular hydrophobic interactions. Kinetic results indicate that early events of protein folding are accelerated in mutants.

• Journal of Radiation Industry
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• v.11 no.3
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• pp.145-149
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• 2017

The cel7A sequence variation was analyzed between the wild type (Lentinula edodes KACC42378) and its cellulase activity enhanced mutant LER277. LER277 was induced by using gamma ray radiation ($^{60}Co$) at the $LD_{99}$ dose (0.94 kGy). Cloning and sequencing results showed that the cel7A coding DNA sequence (CDS) of LER277 had five nucleotide substitutions ($T{\rightarrow}C$, 201, 285 and 744 nt; $A{\rightarrow}G$, 525 nt; $C{\rightarrow}T$, 540 nt) and one hexanucleotide repeat insertion (GGCACC, within 1375-1392 nt) compared to that of the wild type. The Five nucleotide substitutions did not change the deduced amino acids and the hexanucleotide insertion elongated the GT repeat in a serine/threonine/glycine-rich linker. These results suggest that the enhancement of the cellulase activity in LER277 partly stemmed from cel7A changes by which the GT repeat of the linker is elongated.

• Journal of Microbiology and Biotechnology
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• v.7 no.2
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• pp.101-106
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• 1997

Strain improvement for the enhanced production of streptokinase and streptodornase in Streptococcus sp. ATCC 12449 was performed. Strain UB111, a hyperproductive mutant which was isolated by use of nitrosoguanidine and selection of colonies with large clear zones on DNase test agar plates supplemented with $1{\%}$ glucose and $0.5{\%}$ ammonium chloride, produced about 3 fold more streptokinase and streptodornase than the wild type when tested in shake flask fermentations. The enhanced production of both streptokinase and streptodornase was achieved by cultivating the mutant in a pH-controlled fermentor containing fermentation medium enriched with yeast extract ($2.1{\%}$). Under these conditions, the mutant produced 7300 units/ml of streptokinase and 800 units/ml of streptodornase.

• Microbiology and Biotechnology Letters
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• v.31 no.4
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• pp.435-438
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• 2003

Iron required by all organisms is related with diverse biological processes. Most eukaryotes need extra iron to maintain their nutrition balance. However, extra iron supplement gives many problem to solubility in the cells. To increase the bio-availability of iron in cells, yeast was applied to carry the iron with solubility. Selection of yeast mutants with enhanced iron uptake were performed by mutagenesis using the alkylation agent EMS. Eleven mutant strains with enhanced iron uptake were selected by the measurement of iron content with atomic absorption spectrometer. The iron content in mutants was 1.5- to 2.5-fold more than that in wild-type. These mutants could be served as iron-fortified nutrients for food and feed.

• The Korean Journal of Physiology and Pharmacology
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• v.1 no.1
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• pp.19-25
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• 1997

The C-terminus ends of the second putative transmembrane domains of both $M_1$ and $M_2$ muscarinic receptors contain a triplet of amino acid residues consisting of leucine (L), tyrosine (Y) and threonine (T). This triplet is repeated as LYT-TYL in $M_1$ receptors at the interface between the second transmembrane domain and the first extracellular loop. Interestingly, however, it is repeated in a transposedfashion (LYT-LYT) in the sequence of $M_2$ receptors. In our previous work, we investigated the possible significance of this unique sequence diversity for determining the distinct differential receptor function at the two receptor subtypes. However, we found mutation of the LYTTYL sequence of $M_1$ receptors to the corresponding $M_2$ receptor LYTLYT sequence demonstrated markedly enhanced the stimulation of phosphoinositide (PI) hydrolysis by carbachol without a change in its coupling to increased cyclic AMP formation. In this work, thus, the enhanced stimulation of PI hydrolysis in the LYTLYT $M_1$ receptor mutant was further investigated. The stimulation of PI hydrolysis by carbachol was enhanced in the mutant $M_1$ receptor, and this change was not due to alterations in the rate of receptor desensitization or sequestration. The observed larger response to carbachol at mutant $M_1$ receptors was also not due to an artifact resulting from selection of CHO cells which express higher levels of G-proteins or phospholipase C. Our data suggest that although the LYTTYL sequence in $M_1$ muscarinic receptors is not involved in determining receptor pharmacology, mutation of the sequence enhanced the coupling of $M_1$ receptors to the stimulation of phospholipase C.