• Journal of Marine Bioscience and Biotechnology
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• v.2 no.2
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• pp.123-125
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• 2007

Vibrio anguillarum is a gram-negative bacterium that causes vibriosis in approximately 80 different fish species. V. anguillarum produces several exotoxins are correlated with the pathogenesis of vibriosis. This study is focused on the composition of the outer membrane vesicle. Most of gram-negative bacteria produce outer membrane vesicle (OMV) during cell growth. OMV was formed from the outer membrane surface of cell and than released to extracellular environment. OMV consists of outer membrane lipids, outer membrane protein (OMP), LPS, and soluble periplasmic components. Also, they contain toxins, adhesions, and immunomodulatory. Many gram-negative bacteria were studied out forming OMV. In Vibrio sp., formation of OMV by electron microscopy has been reported from V. cholerae and V. parahaemolyticus. In present study, we isolated OMV from V. anguillarum and OMV protein was separated by SDS-PAGE. Magor band was sliced and analyzed by MALDI-TOF. The major protein band of 38kDa was identified as OmpU by MALDI-TOF MS analysis.

• Journal of Microbiology and Biotechnology
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• v.26 no.8
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• pp.1343-1347
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• 2016

Outer membrane vesicles (OMVs) are spherical nanostructures that are ubiquitously shed from gram-negative bacteria both in vitro and in vivo. Recent findings revealed that OMVs, which contain diverse components derived from the parent bacterium, play an important role in communication with neighboring bacteria and the environment. Furthermore, nanoscale proteoliposomes decorated with pathogen-associated molecules attract considerable attention as a non-replicative carrier for vaccines and drug materials. This review introduces recent advances in OMV biogenesis and discusses the roles of OMVs in the context of bacterial communication and virulence regulation. It also describes the remarkable accomplishments in OMV engineering for diverse therapeutic applications.

• BMB Reports
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• v.57 no.8
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• pp.369-374
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• 2024

Antigen 43 (Ag43) proteins, found on the outer membrane of Escherichia coli, are β-sheets that fold into a unique cylindrical structure known as a β-barrel. There are several known structural similarities between bacterial Ag43 autotransporters and physical components; however, the factors that stabilize the barrel and the mechanism for Ag43 passenger domain-mediated translocation across the pore of the β-barrel remain unclear. In this study, we analyzed Ag43β-enhanced green fluorescent protein chimeric variants to provide new insights into the autotransporter Ag43β-barrel assembly, focusing on the impact of the α-helical linker domain. Among the chimeric variants, Ag43β700 showed the highest surface display, which was confirmed through extracellular protease digestion, flow cytometry, and an evaluation of outer membrane vesicles (OMVs). The Ag43β700 module offered reliable information on stable barrel folding and chimera expression at the exterior of the OMVs.

• The Korean Journal of Pharmacology
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• v.28 no.1
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• pp.103-114
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• 1992

Selective quenching of 1,6-diphenyl-1,3,5-hexatriene (DPH) by trinitrophenyl groups was utilized to examine the transbilayer fluidity domains of the model membranes of total lipids (SPMVTL) and phospholipids (SPMVPL) extracted from synaptosomal plasma membrane vesicles. At $37^{\circ}C$, all anisotropy (r), limiting anisotropy $(r_{\infty})$, and order parameter (S) values of DPH in the SPMVTL were larger than those in SPMVPL. The anisotropy, limiting anisotropy, and order parameter of DPH in the inner monolayer were 0.025, 0.033, and 0.070, respectively, greater than calculated for the outer monolayer of SPMVTL. In SPMVPL, the anisotropy, limiting anisotropy, and order parameter of DPH in the inner monolayer were 0.014, 0.018, and 0.047, respectively, greater than calculated for the outer monolayer. Selective quenching of DPH by trinitrophenyl groups was also utilized to examine the effects of barbiturates on the transbilayer fluidity domains of SPMVTL and SPMVPL. Barbiturates did not affect the anisotropy of DPH in the transbilayer domains of SPMVTL. In contrast, barbiturates increased the fluorescence anisotropy, limiting anisotropy, and order parameter of DPH in the SPMVPL in a dose-dependent manner. Barbiturates showed a greater ordering effect on the outer monolayer as compared to the inner monolayer of SPMVPL. Hence, it has been demonstrated for the first time that the Sheetz-Singer hypothesis (1974) may be valid for phospholipid model membranes.

• The Korean Journal of Pharmacology
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• v.28 no.2
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• pp.191-199
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• 1992

Selective quenching of 1,6-diphenyl-1,3,5-hexatriene (DPH) by trinitrophenyl groups was utilized to examine the transbilayer fluidity asymmetry of model membranes of total lipids (SPMVTL) extracted from synaptosomal plasma membrane vesicles (SPMV). The polarization (P), anisotropy (r), limiting anisotropy $(r_{\infty})$, and order parameter (S) of DPH in the inner monolayer were 0.031, 0.025, 0.033, and 0.070, respectively, greater than calculated for the outer monolayer of SPMVTL. Selective quenching of DPH by trinitrophenyl groups was also utilized to examine the effects of n-alkanols on the individual monolayer structure of SPMVTL. n-Alkanols fluidized the hydrocarbon region of bulk SPMVTL, and the potencies of n-alkanols up to 1-nonanol increased with carbon chain length. It appears that the potencies in bilayer fluidization increase by 1 order of magnitude as the carbon chain length increases by two carbon atoms. The cut-off phenomenon was reached at 1-decanol, where further increase in hydrocarbon length resulted in a decrease in pharmacological activity. The n-alkanols had greater fluidizing effects on the outer monolayer as compared to the inner monolayer of SPMVTL, even though these selective effects tended to become weaker as carbon chain length increased. Thus, it has been proven that n-alkanols exhibit selective rather than nonselective fluidizing effects within transbilayer domains of SPMVTL.

• The Korean Journal of Pain
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• v.6 no.2
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• pp.237-246
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• 1993

세포막에서 마취제의 작용점을 규명하기 위하여, 마취제의 많은 부분을 차지하는 n-Alkanol을 이용하여, 소의 synaptosomal plasma membrane vesicles(SPMV)에서 n-Alkanol의 침투 정도를 형광 probe를 이용한 형광소광법을 통하여 검색하였다. n-Alkanols는 SPMV 외부 단층(outer monolayer)의 표면에 주로 분포하되 그 탄소수에 비례하여 소수성 부위에 분포되는 양이 증가되는 경향을 나타내었다(1-decanol은 제외). Methanol, Ethanol, 1-propanol, 1-butanol, 1-pentano, 1-hexanol, 1-heptanol, 1-octanol, 1-nonanol 및 1-decanil은 SPMV 외부 단층의 표면(친수성 부위)에 분포되는 것에 비하여 각각 949, 416.8, 214.8, 90.3, 53.7, 15.20, 6.80, 2.00, 1.03 및 2.40 배가 된다는 것을 확인하였다. 1-decanol은 $C_{10}$인데도 불구하고 $C_8$인 1-octanol에 비하여 적은 양이 소수성 부위에 침투 분포한다는 것이 확인되었다.

• Journal of Microbiology and Biotechnology
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• v.19 no.10
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• pp.1271-1279
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• 2009

To develop low endotoxic and multi-immunogenic outer membrane vesicles (OMVs), a deletion mutant of the msbB gene in Salmonella enterica serovar Typhimurium (S. Typhimurium) was used as a source of low endotoxic OMV, and an expression vector of the canine parvovirus (CPV) VP2 epitope fused to the bacterial OmpA protein was constructed and transformed into the Salmonella ${\Delta}msbB$ mutant. In a lethality test, BALB/c mice injected intraperitoneally with the Salmonella ${\Delta}msbB$ mutant survived for 7 days, whereas mice injected intraperitoneally with the wild type survived for 3 days. Moreover, all mice inoculated orally with the ${\Delta}msbB$ mutant survived for 30 days, but 80% of mice inoculated orally with the wild type survived. The OmpA::CPV VP2 epitope fusion protein was expressed successfully and associated with the outer membrane and OMV fractions from the mutant S. Typhimurium transformed with the fusion protein-expressing vector. In immunogenicity tests, sera obtained from the mice immunized with either the Salmonella msbB mutant or its OMVs containing the OmpA::CPV VP2 epitope showed bactericidal activities against wild-type S. Typhimurium and contained specific antibodies to the CPV VP2 epitope. In the hemagglutination inhibition (HI) assay as a measurement of CPV-neutralizing activity in the immune sera, there was an 8-fold increase of HI titer in the OMV-immunized group compared with the control. These results suggested that the CPV-neutralizing antibody response was raised by immunization with OMV containing the OmpA::CPV VP2 epitope, as well as the protective immune response against S. Typhimurium in BALB/c mice.

• Microbiology and Biotechnology Letters
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• v.27 no.6
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• pp.440-445
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• 1999

In order to investigate structure-antibiotic activity relationships of brevinin-1 and thanatin containing Rana box composed of basic loop formed by disulfide bridge in their arboxy terminus, thanatin, brevinin 1 and their analogues (T-B1, T-B2 and B-T) in which their Rana box sequence exchanged was designed and synthesized by the solid phase method using Fmoc-chemistry. The basic sequence of Rana box of thanatin had more significant effect on both antibacterial and antifungal activity than that of brevinin 1. The tail sequence (QRM) of thanatin was found to be important in its antibacterial and antifungal activity. Rana box sequence of brevinin-1 did not have a significant effect on its antitumor and phospholipid vesicle-aggregating activities. Brevinin-1 showed stronger $\alpha$-helical structure in the membrane-mimicking environment such as SDS micelle than thanatin. A remarkable increase in a-helicity of bervinin-1 plays more important role in antibiotic activity than that of thanatin. Furthermore, antibacterial activity of thanatin against E. coli resulted from the disruptive effect against the outer cell membrane of E. coli.

• Journal of Oral Medicine and Pain
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• v.24 no.4
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• pp.361-374
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• 1999

F. nucleatum is a gram-negative obligate anaerobe which is the principal and most frequent cause of gingival inflammation and is the predominant pathogen isolated in subsequent periodontal breakdown. It is also one of the most numerous bacteria found in subgingival plaque samples from healthy sites; its numbers are about 10-fold greater in plaque from periodontally diseased sites. The purpose of this study is to examine the effects of outer membrane(OM), outer membrane vesicle(OMV), and lipopolysaccharide(LPS) from F. nucleatum ATCC 25586 strain on the growth of human gingival fibroblasts and HOS 941 cells, and on the $TNF-{\alpha}$ production / $TNF-{\alpha}$ mRNA expression of mouse splenocytes. For the examination of cytotoxic effects, $TNF-{\alpha}$ production and $TNF-{\alpha}$ mRNA expression, the MTT assay, the ELISA and the RT-PCR were performed, respectively. All extracts of F. nucleatum tested were cytotoxic to both of human gingival fibroblasts and HOS 941 cells, and the significant difference of cytotoxic activity among the extracts was not observed. In the effects of these extracts on the $TNF-{\alpha}$ production / $TNF-{\alpha}$ mRNA expression of mouse splenocytes, all extracts of F. nucleatum tested also stimulated the $TNF-{\alpha}$ production / $TNF-{\alpha}$ mRNA expression, but the effects of the OM extracts on the $TNF-{\alpha}$ production / $TNF-{\alpha}$ mRNA expression were higher than those of the OMV and the LPS extracts. The pattern of the $TNF-{\alpha}$ mRNA expression was similar to that of the $TNF-{\alpha}$ production. These results indicate that F. nucleatum seems to contribute to the pathogenesis of periodontal diseases at least by its cytotoxicity, directly and its $TNF-{\alpha}$ production, indirectly.

• The Korean Journal of Physiology and Pharmacology
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• v.3 no.5
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• pp.513-519
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• 1999

Direct exposure of renal tubular brush-border membranes (BBM) to free cadmium (Cd) causes a reduction in phosphate (Pi) transport capacity. Biochemical mechanism of this reduction was investigated in the present study. Renal proximal tubular brush-border membrane vesicles (BBMV) were isolated from rabbit kidney outer cortex by Mg precipitation method. Vesicles were exposed to $50{\sim}200\;{\mu}M\;CdCl_2$ for 30 min, then the phosphate transporter activity was determined. The range of Cd concentration employed in this study was comparable to that of the unbound Cd documented in renal cortical tissues of Cd-exposed animals at the time of onset of renal dysfunction. The rate of sodium-dependent phosphate transport $(Na^+-Pi\;cotransport)$ by BBMV was determined by $^{32}P-Iabeled$ inorganic phosphate uptake, and the number of $Na^+-Pi$ cotransporters in the BBM was assessed by Pi-protectable $^{14}C-labeled$ phosphonoformic acid $([^{14}C]PFA)$ binding. The exposure of BBMV to Cd decreased the $Na^+-Pi$ cotransport activity in proportion to the Cd concentration in the preincubation medium, but it showed no apparent effect on the Pi-protectable PFA binding. These results indicate that an interaction of renal BBM with free Cd induces a reduction in $Na^+-Pi$ cotransport activity without altering the carrier density in the membrane. This, in turn, suggest that the suppression of phosphate transport capacity $(V_{max})$ observed in Cd-treated renal BBM is due to a reduction in $Na^+-Pi$ translocation by existing carriers, possibly by Cd-induced fall in membrane fluidity.