• Proceedings of the Korean Society for Applied Microbiology Conference
• /
• 2000.04a
• /
• pp.168-171
• /
• 2000

The thermal stability and catalytic activity of phospholipase A$_1$ from Serratia sp. MK1 were improved by an evolutionary molecular engineering. Two thermostable mutants were isolated after sequential rounds of error-prone PCR to introduce random mutations and filter-based screening of the resultant mutant library, and identified as having six (mutant TA3) and seven (mutant TA13) amino acid substitutions, respectively. Different types of the substitutions were found in two mutants, resulting in the increase of nonploar residues (mutant TA3) or changes between side chains within polar or charged residues (mutant TA13). The wild-type and mutant enzymes were purified, and the effect of temperature on their stability and catalytic activity was investigated. The T$\sub$m/ values of TA3 and TA13 were increased by 7 and 11$^{\circ}C$, respectively. Thus, evolutionary molecular engineering was found to be an effective and efficient approach to increasing thermostability without compromising enzyme activity.

• Journal of Integrative Natural Science
• /
• v.1 no.1
• /
• pp.47-53
• /
• 2008

In a previous study, we showed that Cecropin A (1-8)-Magainin 2 (1-12) hybrid peptide (CA-MA)'s analogue, Anal P5, exhibit broad-spectrum antimicrobial activity. Anal P5, designed by flexible region (positions 9, 10)-substitution, Lys- (positions 4, 8, 14, 15) and Leu- (positions 5, 6, 12, 13, 16, 17, 20) substitutions, showed an enhanced antimicrobial and antitumor activity without hemolysis. The primary objective of the present study was to gain insight into the relevant mechanisms of antimicrobial activities of Anal P5 by using flow cytometric analysis. Anal P5 exhibits strong antifungal activity in a salt concentration independent manner. In addition, Anal P5 causes significant morphological alterations of the bacterial surfaces as shown by scanning electron microscopy, supporting its antibacterial activity. Its potent antibiotic activity suggests that Anal P5 is an excellent candidate as a lead compound for the development of novel antibiotic agents.

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

• Bulletin of the Korean Chemical Society
• /
• v.29 no.9
• /
• pp.1835-1838
• /
• 2008

• Proceedings of the Materials Research Society of Korea Conference
• /
• 1994.05a
• /
• pp.30-30
• /
• 1994

• Journal of the Korean Magnetic Resonance Society
• /
• v.13 no.1
• /
• pp.27-34
• /
• 2009

HP (2-20), a 19-residue peptide derived from the N-terminus of Helicobacter pylori Ribosomal Protein L1, has antimicrobial activity but is not cytotoxic to human erythrocytes. Previously, we have synthesized several analogue peptides to investigate the effects of substitutions on the structure and antimicrobial activity. Substitution of $Gln^{16}$ and $Asp^{18}$ with Trp (Anal 3) caused a dramatic increase in bacterial and fungal lytic activities. In this study, analogue peptides were synthesized to investigate the effects of substitution of Gin and Asp with Phe (Anal 6) or Tyr (Anal 7) in HP (2-20) on its structure and antimicrobial activity. Substitution of Gin and Asp with hydrophobic aromatic residues at position 16 and 18 of HP (2-20) caused increase in antibiotic activity without hemolytic effect. Substitution of Gin and Asp with Trp and Try increased antibiotic activity of HP (220) twice more compared to substitution with Phe. The tertiary structures of Anal 6 and Anal 7 in SDS micelles has been investigated using NMR spectroscopy. The structures revealed that substitutions of the aromatic residues at C-terminus resulted in longer and well defined alpha-helix and improved their antibacterial activities

• Electrical & Electronic Materials
• /
• v.11 no.11
• /
• pp.9-17
• /
• 1998

The dependence of the ionic conductivity on the B-site ion substitution in (Li0.5La0.5)Ti1-xMxO3 (M=Sn, Zr, Mn, Ge) system has been studied. Same valence state and various electronic configuration and ionic radius of Sn4+, Zr4+, Mn4+ and Ge4+(4d10(0.69$\AA$), 4p6(0.72$\AA$), 3d10(0.54$\AA$) and 3d3(0.54$\AA$), respectively) induced the various crystallographic variaton with substitutions. So it was possibleto investigate the crystallographic factor which influence the ionic conduction by observing the dependence of the conductivity on the crystallographic factor which influence the ionic conduction by observing the dependence of the conductivity on the crystallographic variations. We found that the conductivity increased with decreasing the radii of B-site ions or vice versa and octahedron distortion disturb the ion conduction. The reason for this reciprocal proportion of conductivity on the radius of B-site ions has been examined on the base of the interatomic bond strength change due to the cation substitutions. The results were good in agreement with the experimental results. Therefore it could be concluded that the interatomic bond strength change due to the cation substitutions may be the one of major factors influencing the lithium ion conductivity in perovskite(Li0.5La0.5) TiO3system.

• Korean Journal of Breeding Science
• /
• v.42 no.1
• /
• pp.1-10
• /
• 2010

Five mutants were investigated at the molecular level to determine the factors responsible for mutated endosperm types. They were classified as high (HA) or low amylose (LA) phenotypes based on the amylose content in endosperm. The five were previously produced from Ilpum and Shindongjin cultivar treated with N-methyl-N-nitrosourea and gamma-ray irradiation, respectively. Analysis of the genomic structure and expression of Granule-bounded Starch Synthase I (GBSSI) genes revealed that mutants generally showed a higher incidence of nucleotide transition than transversion, and the $A:T{\rightarrow}G:C$ transition was particularly prevalent. The rates of nucleotide substitution in HA mutants were generally higher than those in the LA mutants, leading to higher substitutions of amino acid in the HA mutants. Neither nucleotide substitutions interfering with intron splicing or causing early termination of protein translation were found, nor any large-sized deletions or additions were found in all the mutants. In principle, amylose content can be regulated by three factors: internal alterations of GBSSI protein, the strength of gene expression, and other unknown external factors. Our results indicate that the endosperm mutants from Shindongjin arose from internal alterations of GBSSI proteins, which may be the result of amino acid substitutions. On the other hand, the Ilpum mutants might be principally caused by the alteration of gene expression level. Analysis of another three glutinous cultivars revealed that the major factor leading to glutinous phenotypes is the 23-bp duplicative motif (5'-ACGGGTTCCAGGGCCTCAAGCCC-3') commonly found in exon 2, which results in the premature termination of protein translation leading to the production of a non-functional GBSSI enzyme.

• Molecules and Cells
• /
• v.19 no.1
• /
• pp.114-123
• /
• 2005

Nitropyrene, the predominant nitropolycyclic hydrocarbon found in diesel exhaust, is a mutagenic and tumorigenic environmental pollutant that requires metabolic activation via nitroreduction and ring oxidation. In order to determine the role of ring oxidation in the mutagenicity of 1-nitropyrene, its oxidative metabolites, 1-nitropyrene 4,5-oxide and 1-nitropyrene 9,10-oxide, were synthesized and their mutation spectra were determined in the coding region of hprt gene of CHO cells by a PCR amplification of reverse-transcribed hprt mRNA, followed by a DNA sequence analysis. A comparison of the two metabolites for mutation frequencies showed that 1-nitropyrene 9,10-oxide was 2-times higher than 1-nitropyrene 4,5-oxide. The mutation spectrum for 1-nitropyrene 4,5-oxide was base substitutions (33/49), one base deletions (11/49) and exon deletions (5/49). In the case of 1-nitropyrene 9,10-oxide, base substitutions (27/50), one base deletions (15/50), and exon deletions (8/50) were observed. Base substitutions were distributed randomly throughout the hprt gene. The majority of the base substitutions in mutant from 1-nitropyrene 4,5-oxide treated cells were $A{\rightarrow}G$ transition (15/33) and $G{\rightarrow}A$ transition (8/33). The predominant base substitution, $A{\rightarrow}G$ transition (11/27) and $G{\rightarrow}A$ transition (8/27), were also observed in mutant from 1-nitropyrene 9,10-oxide treated cells. The mutation at the site of adenine and guanine was consistent with the previous results, where the sites of DNA adduct formed by these compounds were predominant at the sites of purines. A comparison of the mutational patterns between 1-nitropyrene 4,5-oxide and 1-nitropyrene 9,10-oxide showed that there were no significant differences in the overall mutational spectrum. These results indicate that each oxidative metabolite exhibits an equal contribution to the mutagenicity of 1-nitropyrene, and ring oxidation of 1-nitropyrene is an important metabolic pathway to the formation of significant lethal DNA lesions.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.28 no.5
• /
• pp.649-658
• /
• 1995

In order to estimate the level of genetic differences among the pleuronectid species, mitochondrial DNAs were isolated from four species: brown sole, Limanda herensteini; marbled sole, Limanda Yokohamae; stone flounder, Kareius bicoloratus; starry flounder, Platichthys stellatus, and the number of nucleotide substitutions was calculated by the restriction fragment length polymorphisms (RFIPs) generated by f4 sin base recognition restriction endonucleases. Total lengths of the mitochondrial DNA were measured as about 17.6 kbp in all species. Ten different composite genotypes were observed in brown sole, four different genotypes in marbled sole, and two different genotypes in starry flounder. However, only one genotype was observed in stone flounder. The calculated haplotypic diversity value of brown sole was higher than that of marbled sole. The average number of nucleotide substitutions per sites in four species was estimated to be 0.0045 in the intraspecies, 0.0344 in the interspecies, and 0.0457 in the genera, respectively. From these results, we could estimate that the genetic differences among interspecies were not influenced by nucleotide substitutions but genetical discontinuous.