• Korean Journal of Microbiology
• /
• v.43 no.4
• /
• pp.325-330
• /
• 2007

RNase E is an essential Escherichia coli endoribonuclease that plays a major role in the decay and processing of a large fraction of RNAs in the cell and expression of N-terminal domain consisted of 1-498 amino acids (N-Rne) is sufficient to support normal cellular growth. By utilizing these properties of RNase E, we developed a genetic system to screen for amino acid substitutions in the catalytic domain of the protein (N-Rne) that lead to various phenotypes. Using this system, we identified three kinds of mutants. A mutant N-Rne containing amino acid substitution in the S1 domain (I6T) of the protein was not able to support survival of E. coli cells, and another mutant N-Rne with amino acid substitution at the position 488 (R488C) in the small domain enabled N-Rne to have an elevated ribonucleolytic activity, while amino acid substitution in the DNase I domain (N305D) only enabled N-Rne to support survival of E. roli cells when the mutant N-Rne was over-expressed. Analysis of copy number of ColEl-type plasmid revealed that effects of amino acid substitution on the ability of N-Rne to support cellular growth stemmed from their differential effects on the ribonucleolytic activity of N-Rne in the cell. These results imply that the genetic system developed in this study can be used to isolate mutant RNase E with various phenotypes, which would help to unveil a functional role of each subdomain of the protein in the regulation of RNA stability in E. coli.

• Journal of the Korean Crystal Growth and Crystal Technology
• /
• v.24 no.6
• /
• pp.229-236
• /
• 2014

A stoichiometric mixture of evaporating materials for $MnAl_2S_4$ single crystal thin films was prepared from horizontal electric furnace. To obtain the single crystal thin films, $MnAl_2S_4$ mixed crystal was deposited on thoroughly etched semi-insulating GaAs(100) substrate by the Hot Wall Epitaxy (HWE) system. The source and substrate temperatures were $630^{\circ}C$ and $410^{\circ}C$, respectively. The crystalline structure of the single crystal thin films was investigated by the photoluminescence and double crystal X-ray diffraction (DCXD). The temperature dependence of the energy band gap of the $MnAl_2S_4$ obtained from the absorption spectra was well described by the Varshni's relation, $E_g(T)=3.7920eV-5.2729{\times}10^{-4}eV/K)T^2/(T+786 K)$. In order to explore the applicability as a photoconductive cell, we measured the sensitivity (${\gamma}$), the ratio of photocurrent to dark current (pc/dc), maximum allowable power dissipation (MAPD) and response time. The results indicated that the photoconductive characteristic were the best for the samples annealed in S vapour compare with in Mn, Al, air and vacuum vapour. Then we obtained the sensitivity of 0.93, the value of pc/dc of $1.10{\times}10^7$, the MAPD of 316 mW, and the rise and decay time of 14.8 ms and 12.1 ms, respectively.

• Applied Biological Chemistry
• /
• v.43 no.3
• /
• pp.179-183
• /
• 2000

Satsuma mandarin(Citrus unshiu Marc. var. miyagawa) was stored at $3^{\circ}C$ and 85% relative humidity, and then the changes of firmness, pectin- degrading enzymes activity and other physicochemical properties of citrus fruits during storage were investigated. Firmness of fruits with 2 m probe was decreased quickly from 1,176.8 g-force to 503.6 g-force, and moisture of peel and flesh were decreased from 75.3% to 74.9%, and from 91.8% to 90.7% during maturation, respectively. Decay ratio was increased to 18.75% after 90 days' storage, and after then it was increased rapidly. Weight loss was increased gradually to 24.5% during long-term storage. Firmness with 2 mm probe were decreased from 538.9 g-force to 336.9 g-force gradually during storage. Peel moisture was decreased from 75.8% to 72.6%, and flesh moisture was also decreased gradually from 90.3% to 88.3% during storage. Exopoly-galacturonase activity of peel and flesh were increased from 326.0 units/100 g to 534.9 units/100 g, and from 63.1 units/100 g to 81.0 units/100 g at 90 day's storage, respectively. After then, He enzyme activities were decreased from 394.0 units/100 g and 38.0 units/100 g, respectively. Pectinesterase activity of peel and flesh were increased from $14.4\;{\mu}mol$ to $38.8{\mu}mol$, and from $26.0{\mu}mol$ to $39.0{\mu}mol$ at 60 days' storage, respectively. After then, the enzyme activities were decreased to $6.0{\mu}mol$ and $8.2{\mu}mol$, respectively.

• Nuclear Medicine and Molecular Imaging
• /
• v.40 no.4
• /
• pp.218-227
• /
• 2006

Purpose: The HSV1-tk reporter gene system is the most widely used system because of its advantage that direct monitoring is possible without the introduction of a separate reporter gene in case of HSV1-tk suicide gene therapy. In this study, we investigate the usefulness of the reporter probe (substrate), $9-(4-[^{18}F]Fluoro-3-hydroxymethylbutyl)$guanine ($[^{18}F]FHBG$) for non-invasive reporter gene imaging using PET in HSV1-tk expressing hepatoma model. Materials and Methods: Radiolabeled FHBG was prepared in 8 steps from a commercially available triester. The labeling reaction was carried out by NCA nucleophilic substitution with $K[^{18}F]/K2.2.2.$ in acetonitrile using N2-monomethoxytrityl-9-14-(tosyl)-3-monomethoxytritylmethylbutyl]guanine as a precursor, followed by deprotection with 1 N HCl. Preliminary biological properties of the probe were evaluated with MCA cells and MCA-tk cells transduced with HSV1-tk reporter gene. In vitro uptake and release-out studies of $[^{18}F]FHBG$ were performed, and was analyzed correlation between $[^{18}F]FHBG$ uptake ratio according to increasing numeric count of MCA-tk cells and degree of gene expression. MicroPET scan image was obtained with MCA and MCA-tk tumor bearing Balb/c-nude mouse model. Results: $[^{18}F]FHBG$ was purified by reverse phase semi-HPLC system and collected at around 16-18 min. Radiothemical yield was about 20-25%) (corrected for decay), radiochemical purity was >95% and specific activity was around >55.5 $GBq/{\mu}\;mol$. Specific accumulation of $[^{18}F]FHBG$ was observed in HSV1-tk gene transduced MCA-tk cells but not in MCA cells, and consecutive 1 hour release-out results showed more than 86% of uptaked $[^{18}F]FHBG$ was retained inside of cells. The uptake of $[^{18}F]FHBG$ was showed a highly significant linear correlation ($R^2=0.995$) with increasing percentage of MCA-tk numeric cell count. In microPET scan images, remarkable difference of accumulation was observed for the two type of tumors. Conclusion: $[^{18}F]FHBG$ appears to be a useful as non-invasive PET imaging substrate in HSV1-tk expressing hepatoma model.