• BMB Reports
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• 제42권1호
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• pp.53-58
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• 2009

The methyl-directed mismatch repair (MMR) mechanism has been extensively studied in vitro and in vivo, but one of the difficulties in determining the biological relationships between the MMR-related proteins is the tendency of MutL to self-aggregate. The properties of a stable MutL homologue were investigated using a thermostable MutL (TmL) from Thermotoga maritima MSB8 and whose size exclusion chromatographic and crosslinking analyses were compatible with a dimeric form of TmL. TmL underwent conformational changes in the presence of nucleotides and single-stranded DNA (ssDNA) with ATP binding not requiring ssDNA binding activity of TmL, while ADPnP-stimulated TmL showed a high ssDNA binding affinity. Finally, TmL interacted with the T. maritima MutS (TmS), increasing the affinity of TmS to mismatched DNA base pairs and suggesting that the role of TmL in the formation of a mismatched DNA-TmS complex may be a pivotal observation for the study of the initial MMR system.

• 미생물학회지
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• 제43권4호
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• pp.250-255
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• 2007

Deinococcus radiodurans recA는 이 미생물의 방사선 저항성을 나타내는 표현형에 필수적이며 재조합성 DNA 수선 과정에 관여한다. 이 과정에서 RecA 단백질은DNA와 결합하여 반응의 활성 종인 RecA nucleoprotein 필라멘트를 형성한다. DNA-의존성 ATPase 활성과 함께, RecA 단배질의 외가닥 DNA 혹은 이중가닥 DNA와의 상호작용은 RecA 단백질이 관여하는 반응의 중심과정으로 이에 관한 분석을 시도하였다. D. radiodurans RecA 단배질은 DNA에 결합한 DNA-단백질 복합체만이 ATPase 활성을 나타내므로, ATP (혹은 dATP) 가수분해를 측정함으로써 RecA와 외가닥 DNA와의 상호작용 정도를 분석하였다. D. radiodurans RecA 단백질은 외가닥 DNA의 염기 구성의 이질성에 영향을 받았으며, homopolymer인 poly(dT)와의 상호작용에서 가장 높은 가수분해 활성을 보였다. Homopolymer인 합성 DNA-의존성 ATP 및 dATP의 가수분해는 pH 6.0과 9.0의 범위에서 다소 일정한속도로 일어났으며 최적 pH는 7.0과 7.5 사이였다. 외가닥 DNA-의존성 ATPase 활성은 염의 존재에 영향을 받아 KCl이 존재하면 다소 억제되나, K-glutamate가 존재하면 오히려 촉진되었다. RecA 단백질과 외가닥 DNA의 상호작용을 ATP 가수분해로 분석하였을 때 2 mM 이상의 magnesium 이온이 DNA 결합반응에 필요하였으며, 비교적 넓은 범위의 pH에서 외가닥 DNA와의 결합반응이 일어나며, 이러한 결합반응은 당량적인 비(1:3, RecA protein: DNA nucleotide)로 일어났다.

• 방사선산업학회지
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• 제5권3호
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• pp.243-247
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• 2011

In this study, a convenient method for the selective immobilization of single strand DNA (ssDNA) on a polymer surface was described. A positively charged polyelectrolyte, poly(diallyldimethylammonium chloride) (PDDA), was spin-coated on a tissue culture petridish and the micropatterns of the PDDA were formed by selective ion implantation through a pattern mask. The surface property of the implanted PDDA was investigated by using a surface profiler and FT-IR spectrometer. Cy3-labeled ssDNA was selectively immobilized on the PDDA patterns through ionic interaction and thus, well-defined ssDNA patterns were obtained.

• Bulletin of the Korean Chemical Society
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• 제30권5호
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• pp.1031-1034
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• 2009

Fluorescence of quinolones including norfloxacin, ciprofloxacin and S- and R-ofloxacin is quenched upon association with single and double-stranded DNA (ss- and ds-DNA). The ratios of fluorescence intensity in the presence of DNA to its absent were plotted with respect to the DNA concentration to construct the Stern-Volmer plot. The slope of the Stern-Volmer plot become larger as the temperature is lowered, ensuring that the fluorescence quenching is static process, i.e., the fluorescence is quenched by formation of the non-fluorescent complex between quinolone and DNA. In the static quenching mechanism, the quenching constant which is equivalent to the slope of the Stern-Volmer plot, is considered as the equilibrium constant for the association of quinolones and DNA. From the temperature-dependent equilibrium constant, ${\Delta}H^0\;and\;{\Delta}S^0$ was obtained using the van’t Hoff relation. In general, association of the quinolone with ds- as well as ss-DNA is energetically favorable (an exothermic) process while the entropy change was unfavorable. Due to the steric effect of the substituents, the effect of the quinolone ring is smaller on the ss-DNA compared to ds-DNA.

• BMB Reports
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• 제35권2호
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• pp.194-198
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• 2002

Eukaryotic replication protein A (RPA) is a single-stranded(ss) DNA binding protein with multiple functions in DNA replication, repair, and genetic recombination. The 70-kDa subunit of eukaryotic RPA contains a conserved four cysteine-type zinc-finger motif that has been implicated in the regulation of DNA replication and repair. Recently, we described a novel function for the zinc-finger motif in the regulation of human RPA's ssDNA binding activity through reduction-oxidation (redox). Here, we show that yeast RPA's ssDNA binding activity is regulated by redox potential through its RPA32 and/or RPA14 subunits. Yeast RPA requires a reducing agent, such as dithiothreitol (DTT), for its ssDNA binding activity. Also, under non-reducing conditions, its DNA binding activity decreases 20 fold. In contrast, the RPA 70 subunit does not require DTT for its DNA binding activity and is not affected by the redox condition. These results suggest that all three subunits are required for the regulation of RPA's DNA binding activity through redox potential.

• Archives of Pharmacal Research
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• 제27권11호
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• pp.1161-1167
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• 2004

Adriblastina, a cancerostatic anthracycline antibiotic, causes considerable oxidative damage to DNA molecules. The interaction of this compound with DNA was investigated using Osteryoung square wave stripping voltammetry (OSWSV) and cyclic voltammetry (CV) at an in situ mercury film electrode. It was found that the equilibrium constant of the bonded oxidized form of the drug was 63 times bigger more important than that of the bonded reduced form. Copper forms 1 metal: 2 drug stoichiometry complex which is highly stable compared to ssDNA-drug interaction and consequently inhibited the drug biochemical damaging effects. Copper complex offered sub-nanogram determination of adriblastina in aqueous and urine media.

• 대한환경공학회지
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• 제32권11호
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• pp.1063-1068
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• 2010

본 연구는 유효한 분변오염의 지표 미생물로서 제안되고 있는 E.coli와, 병원성 바이러스에 대한 지표 미생물로 추천되는 $Q{\ss}$ 파지의 UV 및 UV/$H_2O_2$에 대한 내성을 비교하였다. 이 결과에 의거, 병원성 바이러스 관리를 위한 E.coli 규제의 유효성을 고찰하였다. 또한, 병원성 바이러스에 대한 대체 지표 미생물로써, $Q{\ss}$ 파지, T4 및 lambda 파지의 실제 하수 2차 처리 수중에서의 UV 불활성화를 비교하였다. 실험결과, $Q{\ss}$ 파지와 E.coli는 UV에 대해 거의 동등한 내성을 보였으나, UV/$H_2O_2$ 공정에 대해서는 E.coli보다 $Q{\ss}$ 파지가 더 높은 내성을 보였다. 이로부터, $Q{\ss}$ 파지가 모든 병원성 바이러스의 UV 또는 UV/$H_2O_2$에 대한 내성을 대표한다고 할 수는 없을지라도, 지표 미생물로써 E.coli의 불활성화 효과에만 의존하는 것은, 소독공정에 따라서는 바이러스류에 대해 충분한 소독효과를 얻지 못할 수도 있음을 알 수 있었다. 한편, T4 및 lambda 등 DNA 파지와 불활성화를 비교하였을 경우, $Q{\ss}$ 파지가 UV에 대해 더 낮은 내성을 보였다. 따라서, 불활성화에 대한 지표 바이러스로써 $Q{\ss}$ 파지의 이용은 낮은 UV 조사량의 도입에 의해 결과적으로 불충분한 소독효과를 초래할 수도 있을 것 같다. 이 결과는, 병원성 바이러스류에 대해 보다 충분한 불활성화를 달성하기 위해서는 RNA 파지인 $Q{\ss}$보다 T4 및 lambda 등 DNA 파지가 지표 바이러스로써 보다 유효할 수 있음을 보여준다.

• BMB Reports
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• 제44권5호
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• pp.341-346
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• 2011

The single-stranded DNA binding activity of the Escherichia coli RecA protein is crucial for homologous recombination to occur. This and other biochemical activities of ssDNA binding proteins may be affected by various factors. In this study, we analyzed the effect of $CaCl_2$, NaCl and $NH_4NO_3$ salts in combination with the pH and nucleotide cofactor effect on the ssDNA-binding activity of RecA. The studies revealed that, in addition to the inhibitory effect, these salts exert also a stimulatory effect on RecA. These effects occur only under very strict conditions, and the presence or absence and the type of nucleotide cofactor play here a major role. It was observed that in contrast to ATP, ATP${\gamma}$S prevented the inhibitory effect of NaCl and $NH_4NO_3$, even at very high salt concentration. These results indicate that ATP${\gamma}$S most likely stabilizes the structure of RecA required for DNA binding, making it resistant to high salt concentrations.

• Journal of Microbiology and Biotechnology
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• 제33권12호
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• pp.1543-1551
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• 2023

The recently published high-resolution R388 T4SS structure provides exciting new details about the complete complex of T4SS, including the components making up the stalk and arches, numerous symmetry mismatches between regions of the complex, and an intriguing interpretation of the closed stalk and radial symmetry of the inner membrane complex, which is related to pilus biogenesis assembly. However, there are a few unidentified densities in the electron microscopy map and portions of the identified component sequences for which the structure is not yet known. It is also unclear how well this minimized DNA-transporting T4SS predicts the structure of other T4SSs, such as expanded systems and those that transport proteins rather than DNA. In this review, we evaluate what can be inferred from the recent high-resolution structure of the R388 T4SS with respect to the Cag and Dot/Icm systems. These systems were selected because, given what is currently known about these systems, we expect them to present most structural differences compared to the R388 T4SS structure. Furthermore, we discuss bacterial physiology and diversity, the T4SS structures and their variations between different bacterial species. These insights may prove beneficial for researchers who elucidate the structure and functions of T4SS in different bacterial species.

• Journal of Microbiology and Biotechnology
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• 제25권6호
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• pp.771-781
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• 2015

To examine if the molecular chaperone DnaK operon proteins of Streptococcus suis type 2 (SS2) are involved in adhesion to host cells, the abundance values of these proteins from the surface of two SS2 strains of different adhesion capability were compared. Their roles in growth and adhesion to human laryngeal epithelial cell line HEp-2 cells were investigated on SS2 strain HA9801 and its mutants with DnaK operon genes partially knocked-out (PKO mutant) under heat stress. The major difference was that DnaJ was more abundant in strain HA9801 than in strain JX0811. Pretreatment of the bacteria with hyperimmune sera to DnaJ, but not with those to other proteins, could significantly reduce SS2 adhesion to HEp-2 cells. PKO of dnaJ g ene resulted in decreased SS2 growth at 37℃ and 42℃, and reduced its adhesion to HEp-2 cells. The wild-type strain stressed at 42℃ had increased expression of DnaJ on its surface and elevated adhesion to HEp-2 cells, which was also inhibitable by DnaJ specific antiserum. These results indicate that the DnaJ of S. suis type 2 is important not only for thermotolerance but also for adhesion to host cells. Because DnaJ expression is increased upon temperature upshift with increased exposure on the bacterial surface, the febrile conditions of the cases with systemic infections might help facilitate bacterial adhesion to host cells. DnaJ could be one of the potential candidates as a subunit vaccine because of its good immunogenicity.