記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY-BLACKWELL  

Aims: To clarify whether an antibacterial surfactant, cetyltrimethylammonium bromide (CTAB), induces superoxide stress in bacteria, we investigated the generation of superoxide and hydrogen peroxide and expression of soxR, soxS and soxRS regulon genes in Escherichia coli cells with the treatment of CTAB.
Methods and Results: In situ oxidative stress analyses with BES fluorescent probes revealed that generation of both superoxide and hydrogen peroxide were significantly increased with the CTAB treatment at a sublethal concentration in wild-type strain OW6, compared with the CTAB-resistant strain OW66. The activity of manganese-superoxide dismutase (Mn-SOD), a member of the soxRS regulon proteins, was decreased by the CTAB treatment only in strain OW6. Furthermore, quantitative real-time PCR analyses revealed that expression of the soxRS regulon genes was not upregulated, although soxS was upregulated by the CTAB treatment in strain OW6.
Conclusions: Cetyltrimethylammonium bromide treatment led E. coli cells to a generation state of superoxide and hydrogen peroxide. It was also suggested that superoxide generation was caused by inhibiting SoxS function and decreasing Mn-SOD activity.
Significance and Impact of the Study: It was revealed that excess superoxide generation in bacterial cells play a key action of antibacterial surfactants.

DOI： 10.1111/j.1365-2672.2010.04912.x

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：SPRINGER  

The spontaneous antimicrobial surfactant-resistant mutant, Escherichia coli OW66, has been isolated, and its physiological properties have been characterized in our previous paper (Ishikawa et al., J Appl Microbiol 92:261-268, 2002b). This report revealed that strain OW66 had seven mutations in their chromosomal DNA by comparative genomic hybridization microarray, and that their alternative functions were involved in cell resistance to antimicrobial surfactants. These mutations were located in oppB, ydcR, IVR(vacJ-yfdC), rpoN, rpoB, rpoC, and soxR. Furthermore, seven of the single-mutated isogenic strains and seven of the six-mutated isogenic strains were constructed from strains OW6 (NBRC106482) and OW66, respectively, through homologous recombination, and their resistances to an antimicrobial surfactant were measured using the minimum inhibitory concentration method. These results revealed that all six-mutated strains were more sensitive than strain OW66, and that the soxR66 mutation was independently involved in antimicrobial surfactant resistance of E. coli cells. Expression of soxR66 and soxS was increased in both strains OW66 and OW6-soxR66 without the surfactant treatment by the quantitative real time-polymerase chain reaction analysis, compared with strain OW6. Two-dimensional polyacrylamide gel electrophoresis analysis also revealed that some proteins in the soxRS regulon, including Mn-SOD, were overexpressed in both strains OW66 and OW6-soxR66. These results indicate that the soxR66 mutation leads to the constitutive expression of the soxRS regulon, resulting in the acquired resistance of E. coli cells to an antimicrobial surfactant.

DOI： 10.1007/s00253-010-2638-8

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：SOC ANTIBACTERIAL & ANTIFUNGAL AGENTS, JAPAN  

One hundred and seven soil samples were collected from various places In Japan, and their bisphenol-A (BPA, 2,2-bis(4-Hydroxyphenyl)propane) degradative capacities were evaluated. Eighty-five soil samples possessed BPA degradative capacities, and 26 bacterial strains could be isolated as BPA-degrading bacterium. Sequence analysis of their 16S rRNA genes indicated that 22 isolates belonged to proteobacteria groups, and three of four Gram-positive bacterial strains, YA27, NO13, and NO15, were classified as Bacilli. All isolates except strain YA27 completely degraded 115 mu g/mL BPA in L medium but strain YA27 degraded only 50 mu g/mL BPA. Strain YA27 and three Sphingomonas sp. strains could also grow In basal salt media containing BPA as a sole carbon source (BSMB medium). In HPLC analyses, some isolates, including the three Sphingomonas strains, produced some BPA metabolites in their cultures although the others, Including strain YA27, produced no detectable metabolite. Furthermore, the Pseudomonas strains SU1 and SU4 produced some BPA metabolites that were different from the metabolites detected in the degradation of BPA by the S. bisphenolicum strain AO1. These results suggested that all Isolates could be applicable to the bioremediation of BPA-polluted soil and water. Furthermore, we suggest that Bacillus sp. YA27 and Pseudomonas SU1 and SU4 may exhibit novel BPA metabolism pathways that are distinct from that of S. bisphenolicum AO1.

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY-BLACKWELL  

Aims: To clone and characterize the genes bisdA and bisdB, encoding Ferredoxin(bisd) (Fd(bisd)) and cytochrome P450(bisd) (P450(bisd)), respectively, from the bisphenol A (BPA) degrading Sphingomonas bisphenolicum strain AO1.
Methods and Results: The 3.7 kb region containing bisdA and bisdB was cloned by genome walking and colony hybridization. The deduced N-terminal amino acid sequences of bisdA and bisdB were consistent with those of Fd(bisd) and P450(bisd) proteins characterized in our previous report. Two transposase genes, tnpA1 and tnpA2, were also located upstream and downstream of bisdAB. From amino acid sequence analysis, P450(bisd) has two conserved regions corresponding to the oxygen and heme binding regions of the bacterial cytochrome P450 family. Fd(bisd) was similar to putidaredoxin-type [2Fe-2S] ferredoxins. Escherichia coli BL21 (DE3) cells bearing bisdB- and bisdAB-recombinant pET19b were able to degrade BPA. A spontaneous mutant, strain AO1L, which was unable to degrade BPA, was isolated from the stock culture, and it was confirmed that strain AO1L had no bisdAB region.
Conclusions: P450(bisd) monooxygenase sytem, encoded by bisdAB, is one system required for BPA hydroxylation in S. bisphenolicum strain AO1.
Significance and Impact of the Study: Our results indicate that bisdAB are key genes for BPA degradation in S. bisphenolicum strain AO1.

DOI： 10.1111/j.1365-2672.2008.03843.x

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：OXFORD UNIV PRESS  

We previously reported that nicotine protected against tunicamycin (T(m))-induced ER stress-mediated apoptosis, but not thapsigargin (T(g))-induced apoptosis in PC12 cells. In the present study, we report that the expression of glucose-regulated protein 78 (GRP78) was suppressed by nicotine in Tm-treated PC12 cells. Interestingly, the GRP78 expression was not changed by nicotine in Tg-treated cells. Moreover, nicotine reduced the activation of caspase-12 in Tm-treated cells, but not in Tg-treated cells. These results suggest that nicotine prevented Tm-induced ER stress-mediated apoptosis by attenuating an early stage of Tm-induced ER stress. It was possible that the suppression of GRP78 expression by nicotine was achieved through the suppression of the Ire1-XBP1 and/or ATF6 pathways. We observed that nicotine suppressed the Tm-induced, but not Tg-induced, splicing of XBP1 mRNA, and also suppressed the Tm-induced, but not Tg-induced, production of cleaved ATF6 in PC12 cells. These results indicate that the suppression of Ire1-XBP1 and ATF6 pathways contributes to the suppression of GRP78 expression by nicotine in Tm-treated PC12 cells, suggesting that nicotine suppresses a common step upstream of both the Ire1-XBP1 and ATF6 pathways which are required for the expression of GRP78 during Tm-induced ER stress.

DOI： 10.1093/jb/mvn063

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：SPRINGER  

Bisphenol A (2,2-bis(4-hydroxyphenyl) propane, BPA), which is used as a synthetic resin material or a plasticizer, is a pollutant that possesses endocrine-disrupting activity. Bioremediation of BPA is used to decrease its polluting effects, and here we report a novel bacterial strain AO1, which is able to degrade BPA. This strain was isolated using enrichment cultivation from a soil sample from a vegetable-growing field; the sample was one of 500 soil samples collected across Japan. Strain AO1 degraded 100 mg/l BPA to an undetectable level within 6 h in MYPG medium (containing malt extract, yeast extract, peptone, and glucose) and within 48 h in minimum medium containing 1% glucose at 30 degrees C. Strain AO1 can utilize BPA as a sole source of carbon and as an energy source under aerobic conditions. The estrogenic activity of BPA in MYPG medium was ultimately reduced by strain AO1, although the activity initially increased. Taxonomical analysis showed that strain AO1 is closely related to Sphingomonas chlorophenolicum and S. herbicidovorans, neither of which have a capacity for BPA degradation. DNA-DNA hybridization showed that strain AO1 is a novel species of the Sphingomonas genus, and we designated AO1 as S. bisphenolicum.

DOI： 10.1007/s10532-006-9059-5

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Society for Antibacterial and Antifungal Agents Japan  

We investigated characteristics of the corrosion of stainless steel specimens by bacteria and the effects of using antimicrobial coating on the surface for inhibiting corrosion. Bacillus sp. 2-A and Staphylococcus sp. 2-1 cells adhered tightly to a stainless steel SUS304 specimen, formed a microcolony or biofilm, and had highly corrosive activities. Microbially influenced corrosion (MC) was observed under or around adhering cells. However, dead cells were markedly less active than viable cells not only in corroding the specimen but also in adhering to its surface. The culture supernatant was not able to induce the corrosion of SUS304 effectively. A protamine coating on the specimen killed bacterial cells only on its surface, interfered with cell adhesion, and inhibited MC. From these results, adhesion of viable cells to the surface of a SUS304 specimen led to the outbreak of MC. Protamine was also found to be an effective substance tested for protecting the specimen from both cell adhesion and surface MC. We suggest that a protamine coating can be applied as a convenient and inexpensive corrosion prevention method.

DOI： 10.4265/bio.12.21

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER SOC MICROBIOLOGY  

In a previous study (M. Sasaki, J. Maki, K. Oshiman, Y. Matsumura, and T. Tsuchido, Biodegradation 16:449-459, 2005), the cytochrome P450 monooxygenase system was shown to be involved in bisphenol A (BPA) degradation by Sphingomonas sp. strain AO1. In the present investigation, we purified the components of this monooxygenase, cytochrome P450 (P450(bisd)), ferredoxin (Fd(bisd)), and ferredoxin reductase (Red(bisd)). We demonstrated that P450(bisd) and Fd(bisd) are homodimeric proteins with molecular masses of 102.3 and 19.1 kDa, respectively, by gel filtration chromatography analysis. Spectroscopic analysis of Fd(bisd) revealed the presence of a putidaredoxin-type [2Fe-2S] cluster. P450(bisd), in the presence of Fd(bisd), Red(bisd), and NADH, was able to convert BPA. The K-m and k(cat) values for BPA degradation were 85 +/- 4.7 mu M and 3.9 +/- 0.04 min(-1), respectively. NADPH, spinach ferredoxin, and spinach ferredoxin reductase resulted in weak monooxygenase activity. These results indicated that the electron transport system of P450(bisd) might exhibit strict specificity. Two BPA degradation products of the P450(bisd) system were detected by high-performance liquid chromatography analysis and were thought to be 1,2-bis(4-hydroxyphenyl)-2-propanol and 2,2-bis(4-hydroxyphenyl)-1-propanol based on mass spectrometry-mass spectrometry analysis. This is the first report demonstrating that the cytochrome P450 monooxygenase system in bacteria is involved in BPA degradation.

DOI： 10.1128/AEM.71.12.8024-8030.2005

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：SPRINGER  

The capacity and pathway of bisphenol A [BPA; 2,2- bis(4-hydroxyphenyl) propane] degradation in Sphingomonas sp. strain AO1, which was isolated from the soil of a vegetable-growing field in Japan, were investigated. The bacterial strain was able to grow in a basal mineral salt medium containing BPA as the sole carbon source ( BSMB medium), and was able to degrade 115 mug ml(-1) BPA in 6 h in L medium. Several BPA metabolites were detected in the culture supernatant by HPLC and then identified by GC-MS and LC-MS-MS. These compounds were confirmed to be the same as those reported for other BPA-degrading bacteria. BPA degradation by cells in the basal mineral salt medium was induced by BPA, and activity was detected only in the intracellular soluble fraction in the presence of coenzymes, such as NADH, NAD+, NADPH or NADP+. The addition of metyrapone, a cytochrome P450 inhibitor, to BSMB medium resulted in a decrease in BPA degradation and cell growth. The BPA-degradation activity of the intracellular soluble fraction was also inhibited by the cytochrome P450 inhibitor. Carbon monoxide difference spectra indicated that cytochrome P450 was present in the cells and that the amount of cytochrome P450 corresponded to the cellular BPA-degradation activity. Our results provide evidence that the cytochrome P450 system is involved in BPA metabolism in Sphingomonas sp. strain AO1.

DOI： 10.1007/s10532-004-5023-4

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記述言語：日本語   出版者・発行元：The Visualization Society of Japan  

In the field of biotechnology at present, in order to evaluate the viability of bacteria, colony counting is generally used. However, colony counting requires 10 to 48 hours, so delicate changes with passage of time cannot be measured. In this study, we developed a measurement tool of bacterial motion by applying PIV technique in order to evaluate the viability of bacteria quickly. By this tool, not only translational motion but also rotational motion velocity of bacteria can be measured. So the viability of wild strains with tumbling can be evaluated without mutants.

DOI： 10.3154/jvs.23.Supplement1_451

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER SOC MICROBIOLOGY  

The properties of the bactericidal action of silver zeolite as affected by inorganic salts and ion chelators were similar to those of silver nitrate. The results suggest that the contact of the bacterial cell with silver zeolite, the consequent transfer of silver ion to the cell, and the generation of reactive oxygen species in the cell are involved in the bactericidal activity of silver zeolite.

DOI： 10.1128/AEM.69.7.4278-4281.2003

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掲載種別：研究論文（学術雑誌）  

In order to investigate the contribution of the multidrug efflux pumps to cellular resistance to a variety of agents including cationic surfactants, we cloned genes of major drug efflux pumps of Escherichia coil and transformed strain JM109 with plasmids encoding those genes. In emrAB-, mdfA- and emrE-recombinants, the resistance to carbonylcyanide-m-chlorophenylhydrazone (CCCP), chloramphenicol, and paraquat, respectively, was increased compared to their parent bearing pUC18. Also, in mdfA- and emrE-recombinants, the resistance to cetyltrimethylammonium bromide (CTAB) was increased. However, no significant effects of abundant numbers of emrD and envCD could be observed on resistance to agents examined.

DOI： 10.4265/bio.7.115

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：BLACKWELL PUBLISHING LTD  

To examine functions of two small heat shock proteins of Escherichia coli , IbpA and IbpB, we constructed His-IbpA and His-IbpB, in which a polyhistidine tag was fused to the N-terminals. Both purified His-IbpA and His-IbpB formed multimers, which have molecular masses of about 2.0-3.0 MDa and consist of about 100-150 subunits. They suppressed the inactivation of several enzymes including citrate synthase and 6-phosphogluconate dehydrogenase by heat, potassium superoxide, hydrogen peroxide and freeze-thawing, but not the inactivation of glyceraldehyde-3-phosphate dehydrogenase by hydrogen peroxide. Both His-IbpA and His-IbpB suppressed enzyme inactivation by various treatments and were also found to be associated with their non-native forms. However, both His-IbpA and His-IbpB were not able to reactivate enzymes inactivated by heat, oxidants or guanidine hydrochloride. When heated to 50 degreesC, each multimeric form of His-IbpA or His-IbpB was dissociated to form a monomer for His-IbpA, and an oligomer of about one-quarter size for His-IbpB. These structural changes were reversible, as both heated proteins regained the multimeric structures after incubation at 25 degreesC. However, when exposed to hydrogen peroxide or potassium superoxide, the large multimeric forms of His-IbpA and His-IbpB were maintained. The results suggest that His-IbpA and His-IbpB suppress the inactivation of enzymes and bind non-native proteins to protect their structures from heat and oxidants.

DOI： 10.1046/j.1432-1033.2002.02958.x

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：BLACKWELL PUBLISHING LTD  

Aims: In order to clarify the involvement of an energy-yielding system in the antibacterial action of surfactants, the effects of carbon source and anaerobiosis during the growth period on the surfactant sensitivity of Escherichia coli cells were investigated.
Methods and Results: Cetyltrimethylammonium bromide (CTAB) and N -dodecyl-N ,N -dimethylglycine, at relatively low concentrations, caused a delay in growth of E. coli cells. Cells grown in M9 medium supplemented with glycerol, succinate or acetate as a carbon source were more sensitive to surfactants and had a higher respiratory activity than those grown with glucose. Cultivation under anaerobiosis made cells resistant to CTAB.
Conclusions: Bacterial sensitivity to surfactants was affected by carbon source and anaerobiosis.
Significance and Impact of the Study: The results obtained should be helpful in determining suitable conditions of treatment in the practical use of surfactants for bacterial decontamination.

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY-BLACKWELL  

Aims: In order to investigate the mechanism of bacterial resistance to surfactants, a spontaneous mutant of Escherichia coli, OW66, resistant to a cationic surfactant cetyltrimethylammonium bromide (CTAB), was isolated and its physiological properties analysed.
Methods and Results: Strain OW66 grew in M9 medium containing CTAB at 45 mumol l(-1), whereas its parent strain, OW6, did not, even at 15 mumol l(-1). The mutant was also resistant to some other surfactants, antibiotics, heavy metals, organic solvents and oxidants examined. To determine the differences in physiology between strains OW66 and OW6, the compositions of their cell surface structures were analysed. In strain OW66, the relative content of OmpC in particular was higher than that of OmpF, whereas a reverse situation was seen in OW6 strain. The lipopolysaccharide (LPS) profile was different between these strains, and altered LPS in strain OW66 was suggested to be involved in the resistance to CTAB.
Conclusions: A CTAB-resistant E. coli isolate possesses an altered outer membrane.
Significance and Impact of the Study: Treatment with a relatively low concentration of CTAB was found to introduce multi-drug resistance into bacterial cells. This acquired resistance should be taken into account with the frequent use of surfactants in industries and various environments.

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掲載種別：研究論文（学術雑誌）  

In the photoreaction of Escherichia coli IM303 (superoxide dismutase (SOD)-deficient mutant) and MM294 (wild-type strain) with TiO2 particles, the viability of strain IM303 decreased linearly with photoreaction time, while the time profile of the viability of strain MM294 exhibited a curved form. Using strain MM294 with varied initial SOD activities, the TiO2 photoreaction tests were conducted at incident light intensities of I0 = 4,8 and 14W m-2, and the time profiles of bacterial viabilities were analyzed on the basis of the series-event model. The value of n (corresponding to the step number in the series reaction kinetics described by the model) increased with an increase in initial SOD activity (ASOD,0), giving a mean value of ASOD,0/n = 7.1 × 10-9 U cell-1 under the conditions examined. SOD activities in the cells of strain MM294 with ASOD,0 = 1.9 × 10-8 and 4.0 × 10-8 U cell-1 decreased with the progress of photoreaction conducted at I0 = 14W m-2. The transition of intracellular SOD activities expressed was in agreement with the observed data by considering the changes in bacterial cell populations with varied SOD activities based on the proposed model. © 2002 Society of Chemical Industry.

DOI： 10.1002/jctb.619

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：BLACKWELL PUBLISHING LTD  

Aims: In order to clarify the involvement of an energy-yielding system in the antibacterial action of surfactants, the effects of carbon source and anaerobiosis during the growth period on the surfactant sensitivity of Escherichia coli cells were investigated.
Methods and Results: Cetyltrimethylammonium bromide (CTAB) and N -dodecyl-N ,N -dimethylglycine, at relatively low concentrations, caused a delay in growth of E. coli cells. Cells grown in M9 medium supplemented with glycerol, succinate or acetate as a carbon source were more sensitive to surfactants and had a higher respiratory activity than those grown with glucose. Cultivation under anaerobiosis made cells resistant to CTAB.
Conclusions: Bacterial sensitivity to surfactants was affected by carbon source and anaerobiosis.
Significance and Impact of the Study: The results obtained should be helpful in determining suitable conditions of treatment in the practical use of surfactants for bacterial decontamination.

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記述言語：日本語   出版者・発行元：高温学会  

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：OXFORD UNIV PRESS  

To investigate the function of Escherichia coli small heat shock proteins, IbpA and IbpB, we constructed ibpA-, ibpB- and ibpAB-overexpressing strains and also an ibpAB-disrupted strain. The ibpA-, ibpB- and ibpAB-overexpressing strains were found to be resistant not only to heat but also to superoxide stress. However, the ibpAB-disrupted strain was not more sensitive to these stresses than the wild-type strain. The heat sensitivity of a rpoH amber mutant was partially suppressed by the overexpression of plac::ibpAB. These results suggest that IbpA and IbpB may be involved in the resistances to heat and oxidative stress. (C) 2000 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Society for Antibacterial and Antifungal Agents Japan  

We investigated the change in the location of intracellular proteins in Escherichia coli cells after heat treatment at 55°C in a buffer. After heat treatment, the amount of soluble proteins in cell extracts decreased and those of urea-soluble proteins, which were sedimented by high-speed centrifugation and then solubilized with 6 M urea solution, increased correspondingly. Based on the sucrose density gradient ultracentrifugation analysis, two separate peak fractions of the sedimentary intracellular proteins appeared. One was sedimented at the bottom and the other was cosedimented with the membranes. The sedimentary intracellular proteins in a rpoH mutant of E. coli after heat shock to 45°C in a growth medium were also separated in two peak fractions, as in its parent strain heated at 55°C in the buffer, whereas they were hardly detected in the case of the parent strain after the heat-shocked to 45°C. In the case of an IbpAB-overexpressing strain heated to 50°C in a growth medium, sedimentary intracellular proteins did not seem to be cosedimented with the membranes and also protein aggregation decreased, compared with the case of its parent strain heated at 50°C. These results suggest that heat-denatured intracellular proteins not only aggregate but also interact with the membranes and also that at least some of heat shock proteins including IbpAB may be involved in the suppression of protein aggregation.

DOI： 10.4265/bio.5.81

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER SOC MICROBIOLOGY  

We constructed a sodA-disrupted mutant of Bacillus subtilis 168, BK1, by homologous recombination. The mutant was not able to grow in minimal medium without Mn(II). The spore-forming ability of strain BK1 was significantly lower in Mn(II)-depleted medium than that of the wild-type strain. These deleterious effects caused by the sodA mutation were reversed when an excess of Mn(IT) was used to supplement the medium, Moreover, the growth inhibition by superoxide generators in strain BK1 and its parent strain was also reversed by the supplementation with excess Mn(II), We therefore estimated the Mn-dependent superoxide-scavenging activity in BK1 cells. Whereas BK1 cells have no detectable superoxide dismutase (Sod) on native gel, the superoxide-scavenging activity in crude extracts of BK1 cells grown in Mn(II)-supplemented LB medium (10 g of tryptone, 5 g of yeast extract, and 5 g of NaCl per liter) was significantly detected by the modified Sod assay method without using EDTA. The results obtained suggest that Mn, as a free ion or a complex with some cellular component, can catalyze the elimination of superoxide and that both SodA and Mn(II) are involved not only in the superoxide resistance of vegetative cells but also in sporulation.

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER SOC MICROBIOLOGY  

Bacillus subtilis was found to possess one detectable superoxide dismutase (Sod) in both vegetative cells and spores. The Sod activity in vegetative cells was maximal at stationary phase. Manganese was necessary to sustain Sod activity at stationary phase, but paraquat, a superoxide generator, did not induce the expression of Sod, The specific activity of purified Sod was approximately 2,600 U/mg of protein, and the enzyme was a homodimer protein with a molecular mass of approximately 25,000 per monomer. The gene encoding Sod, designated sodA, was cloned by the combination of several PCR methods and the Southern hybridization method. DNA sequence analysis revealed the presence of one open reading frame consisting of 606 bp. Several putative promoter sites were located in the upstream region of sodA. The deduced amino acid sequence showed high homology with other bacterial manganese Sods. Conserved regions in bacterial manganese Sod could also be seen. The phenotype of double mutant Escherichia coli sodA sodB, which could not grow in minimal medium without supplemental amino acids, was complemented by the expression of B. subtilis sodA.

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER CHEMICAL SOC  

The behaviors of heat-induced translocation of cytoplasmic beta-galactosidase to periplasm across the inner membrane of Escherichia coli cells were investigated in order to apply such phenomena to the process for production and separation of intracellular biomolecules. The heat stress was found to induce translocation of cytoplasmic beta-galactosidase (beta-gal) together with reduction of the amounts of intracellular soluble proteins and formation of their inactive aggregates. The translocation of beta-gal was then analyzed using (a) the location factor of beta-gal (LFG), which meant enzyme location in the cells and could be determined from the kinetic analysis of enzyme release process, and (b) the percentage of beta-gal activity in periplasm after solublizing the outer membrane of E. coli cells by lysozyme/EDTA treatment. The LFG values were maximized when cells were stressed at the temperature of 42-47 degrees C. From the results on the surface properties of both beta-gal and cell membrane under the heat stress, it is suggested that (1) the conformational change of cytoplasmic oligomeric beta-gal to the partially dissociated and/or unfolded state with higher local hydrophobicity, (2) the increase in membrane fluidity of inner membrane, (3) the enhancement of hydrophobic interaction between lipid and protein, and (4) the inhibition of its translocation by GroEL restabilizing the proteins could underlie the heat-induced translocation of beta-gal across the inner membrane. The possibility to apply the heat-induced translocation of beta-gal for the enhancement of the target selectivity at the process upstream is finally presented.

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Society for Antibacterial and Antifungal Agents Japan  

The heat resistance of Bacillus subtills 168 spores suspended in several kinds of oil was investigated. In soybean oil, the D value of the thermal death of spores at 98.4°C was 53 min, being about 7 times greater than that in potassium phosphate buffer. The Z value was 71°C in soybean oil but 11°C in the buffer. A similar increase in the D value was observed in other edible oils and n-hexadecane. The differences in the heat resistance of spores among these oils seemed to depend upon the water content of oils even at considerably low levels. Furthermore, the dehydration of oils by dry heat increased the heat resistance of spores. In addition, freeze-dried spores were more resistant to heat than frozen spores in soybean oil. In an open heating system, the spore resistance was higher than that in a closed system. These results suggest that the water content of oil substantially determines the heat resistance of spores suspended in it.

DOI： 10.4265/bio.3.87

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Society for Antibacterial and Antifungal Agents Japan  

For the evaluation of pollutant toxicity in the aquatic environment, a novel principle of a dynamic image analysis of bacterial motion by using a tumbling-ability-defective mutant of Escherichia coli K-12, OW22, isolated as a laboratory indicator organism, is proposed. The average linear motion speed of intact cells of this mutant was approximately 32 μ m/s and decreased markedly by the presence of model chemical pollutants tested. It was suggested that the mutant should be useful as an indicator organism for the evaluation of pollutant toxicity by the dynamic image analysis method.

DOI： 10.4265/bio.1.61

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Society for Antibacterial and Antifungal Agents Japan  

A novel method of killing bacterial cells by cell autolysis induction, 'the suicide induction method,' was proposed. Cold shock treatment and the addition of a surfactant or a high concentration of monovalent cations used for autolysis induction were found to cause the substantial cell death of Bacillus subtilis. Enumeration with the conventional colony counting of the wild-type B. subtilis and its autolysis-defective mutant suggests that the autolysis induction is responsible for the induced cell death, namely bacterial suicide. Although no direct relationship between the cell lysis detected by the optical density decrease and the reduction in colony counts was observed, the evaluation of cell death by the respiratory activity measurement supported the above suggestion. In a continuous tube-flow system consisting of a cooling-warming cycle, bacterial suicide of B. subtilis in substantial numbers was observed. We suggest that 'the suicide induction method' may be applicable for practical control of some bacteria.

DOI： 10.4265/bio.1.19

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：CHAPMAN HALL LTD  

Two types of superoxide dismutase genes, sodA and sodB, were fused to beta-galactosidase gene (lacZ), in order to quantitatively study the effect of oxygen concentration on the gene expression of sodA and sodB. beta-Galactosidase activity derived from the sodA-lacZ fusion was induced by shifting from anaerobic condition to aerobic condition. Maximum activity (9.4 X 10(3) U/OD660) was observed when oxygen partial pressure was. 0.6 atm. On the contrary, gene expression level for the sodB-lacZ gene fusion was about two times higher during anaerobic condition than that during aerobic condition. From these results it was concluded that oxygen positively affected the gene expression of sodA and negatively affected the gene expression of sodB. An inducible expression vector using the sodA regulatory region was also constructed.

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：SOC BIOSCIENCE BIOENGINEERING JAPAN  

Fifty-five oxygen sensitive mutants of Escherichia coli K-12 were isolated by chemical mutagenesis, two of which were manganese superoxide dismutase (Mn-SOD) defective mutants. These mutants (no. 34, no. 58) could grow anaerobically in minimal medium, but not under aerobic conditions. Mn-SOD of no. 58 was induced by adding paraquat or o-phenanthroline. However, the induction level was much lower than that of the wild-type strain without induction, inferring that strain no. 58 is a repressor-overproducing mutant. Strain no. 34 produced a mutant Mn-SOD enzyme (M') which migrated slowly in native PAGE. Protein M' was purified from strain no. 34 and the molecular weight was determined by gel filtration. Protein M' was found to be a dimer of identical subunits, as is the wild-type enzyme. The specific activity of M' is the same as that of the wild type. From these results, it was inferred that the mutant enzyme (M') may be more positively charged or less negatively charged than the wild type and that the productivity of M' was much lower than that of the wild-type enzyme. It was found that multicopies of the ilvD gene complemented the auxotrophic properties of mutant no. 34 under aerobic conditions. Since the gene product (alpha,beta-dihydroxyisovalerate dehydratase) is very susceptible to inactivation by superoxide radicals, a large amount of the enzyme must be needed for strain no. 34 to grow aerobically. In contrast, such a complementary gene was not cloned in strain no. 58.

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記述言語：日本語   出版者・発行元：日本生物工学会  

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記述言語：日本語   出版者・発行元：日本生物工学会  

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記述言語：日本語   出版者・発行元：日本生物工学会  

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記述言語：日本語   出版者・発行元：日本生物工学会  

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記述言語：日本語   出版者・発行元：日本生物工学会  

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記述言語：日本語   出版者・発行元：日本生物工学会  

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記述言語：日本語   出版者・発行元：日本生物工学会  

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記述言語：日本語   出版者・発行元：社団法人日本農芸化学会  

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記述言語：日本語   出版者・発行元：社団法人日本農芸化学会  

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