• Journal of the Korean Society for Industrial and Applied Mathematics
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• 제17권2호
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• pp.87-102
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• 2013

An n-string tangle is a three dimensional ball with n-strings which are properly embedded in the ball. In early 90's, C. Ernst and D. Sumners first used a tangle to describe a DNA-protein complex. In this model, DNA is represented by a string and protein is represented by a ball. Mu is a protein which binds to DNA at three sites and a DNA-Mu complex is called Mu-transpososome. Knowing the DNA topology within Mu-transpososome is very important to understand DNA transposition by Mu protein. In 2002, Pathania et al. determined that the DNA configuration within the Mu transpososome is three branched and five noded [12]. In 2007, Darcy et al. analyzed this by using mathematical tangle and concluded that the three branched and five noded DNA configuration is the only biologically reasonable solution [4]. In this paper, based on the result of Pathania et al. and Darcy et al., the author determines the DNA topology within the DNA-Mu complex after the whole Mu transposition process. Furthermore, a new experiment is designed which can support the Pathania et al.'s result. The result of this new experiment is predicted through mathematical knot thory.

• Molecules and Cells
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• 제42권7호
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• pp.546-556
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• 2019

Protein phosphatase 4 (PP4) is a crucial protein complex that plays an important role in DNA damage response (DDR), including DNA repair, cell cycle arrest and apoptosis. Despite the significance of PP4, the mechanism by which PP4 is regulated remains to be elucidated. Here, we identified a novel PP4 inhibitor, protein phosphatase 4 inhibitory protein (PP4IP) and elucidated its cellular functions. PP4IP-knockout cells were generated using the CRISPR/Cas9 system, and the phosphorylation status of PP4 substrates (H2AX, KAP1, and RPA2) was analyzed. Then we investigated that how PP4IP affects the cellular functions of PP4 by immunoprecipitation, immunofluorescence, and DNA double-strand break (DSB) repair assays. PP4IP interacts with PP4 complex, which is affected by DNA damage and cell cycle progression and decreases the dephosphorylational activity of PP4. Both overexpression and depletion of PP4IP impairs DSB repairs and sensitizes cells to genotoxic stress, suggesting timely inhibition of PP4 to be indispensable for cells in responding to DNA damage. Our results identify a novel inhibitor of PP4 that inhibits PP4-mediated cellular functions and establish the physiological importance of this regulation. In addition, PP4IP might be developed as potential therapeutic reagents for targeting tumors particularly with high level of PP4C expression.

• Journal of Microbiology
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• 제44권5호
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• pp.508-514
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• 2006

In this study, the double-stranded DNA-dependent activities of Deinococcus radiodurans RecA protein (Dr RecA) were characterized. The interactions of the Dr RecA protein with double-stranded DNA were determined, especially dsDNA-dependent ATP hydrolysis by the Dr RecA protein and the DNA strand exchange reaction, in which multiple branch points exist on a single RecA protein-DNA complex. A nucleotide cofactor (ATP or dATP ) was required for the Dr RecA protein binding to duplex DNA. In the presence of dATP, the nucleation step in the binding process occurred more rapidly than in the presence of ATP. Salts inhibited the binding of the Dr RecA protein to double-stranded DNA. Double-stranded DNA-dependent ATPase activities showed a different sensitivity to anion species. Glutamate had only a minimal effect on the double-stranded DNA-dependent ATPase activities, up to a concentration of 0.7 M. In the competition experiment for Dr RecA protein binding, the Dr RecA protein manifested a higher affinity to double-stranded DNA than was observed for single-stranded DNA.

• Animal cells and systems
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• 제10권1호
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• pp.1-6
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• 2006

Previous studies showed that Cdc7 kinase of Schizosaccharomyces pombe phosphorylated the minichromosome maintenance (Mcm) complex efficiently in the presence of spMcm10 protein. The biochemical properties of the phosphorylated Mcm complexes were examined to understand the activation mechanism of the Mcm complex by Cdc7 kinase. The phosphorylation of Mcm complex in the presence of spMcm10 by Cdc7 kinase did not affect the stability of the Mcm complex containing all six subunits, and the changes in the sedimentation properties were not observed after the phosphorylation. The reconstitution of the Mcm complex using the purified proteins showed that the phosphorylation of Mcm2 proteins did not affect the interactions between Mcm proteins. The phosphorylation of the Mcm2-7 complex at the same condition also did not activate the other biochemical activities such as DNA helicase and single stranded (ss) DNA binding activities. On the other hand, spMcm10 protein that was used for the stimulation of Mcm phosphorylation showed single stranded DNA binding activity, and inhibited the DNA helicase activity of the Mcm4/6/7 complex. These inhibitory effects were reduced by the addition of Cdc7 kinase, suggesting that the phosphorylation by Cdc7 kinase decreased the interactions between spMcm10 and the Mcm complex. Taken together, these results suggested that the phosphorylation by Cdc7 kinase alone is not sufficient for the remodeling and the activation of the Mcm complex, and the additional factors or the phosphorylations might be required for the activation of the Mcm complex.

• BMB Reports
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• 제34권6호
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• pp.489-495
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• 2001

DNA damage by UV and environmental agents are the major cause of genomic instability that needs to be repaired, otherwise it give rise to cancer. Accordingly, mammalian cells operate several DNA repair pathways that are not only responsible for identifying various types of DNA damage but also involved in removing DNA damage. In mammals, nucleotide excision repair (NER) machinery is responsible for most, if not all, of the bulky adducts caused by UV and chemical agents. Although most of the proteins involved in NER pathway have been identified, only recently have we begun to gain some insight into the mechanism by which proteins recognize damaged DNA. Binding of Xeroderma pigmentosum group C protein (XPC)-hHR23B complex to damaged DNA is the initial damage recognition step in NER, which leads to the recruitment of XPA and RPA to form a damage recognition complex. Formation of damage recognition complex not only stabilizes low affinity binding of XPA to the damaged DNA, but also induces structural distortion, both of which are likely necessary for the recruitment of TFIIH and two structure-specific endonucleases for dual incision.

• BMB Reports
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• 제33권2호
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• pp.155-161
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• 2000

The saccharomyces cerevisiae Rad27, a structure-specific endonuclease for the okazaski fragment maturation has been known to interact genetically and biochemically with Dna2, an essential enzyme for DNA replication. In an attempt to define the significance of the interaction between the two enzymes, we expressed and purified both Dna2 and Rad27 proteins. In this report, Rad27 could not form a complex with Dna2 in the three different analyses. The analyses included glycerol gradient sedimentation, protein-column chromatography, and coinfection of baculoviruses followed by affinity purification. This is in striking contrast to the previous results that used crude extracts. These results suggest that the interaction between the two proteins is not sufficiently stable or indirect, and thus requires an additional protein(s) in order for Rad27 and Dna2 to form a stable physical complex. This result is consistent with our genetic findings that Schizosaccharomyces pombe Dna2 is capable of interacting with several proteins that include two subunits of polymerase $\delta$, DNA ligase I, as well as Fen-1. In addition, we found that the N-terminal modification of Rad27 abolished its enzymatic activity. Thus, as suspected, we found that on the basis of the structure determination, N-terminal methionine indeed plays an important role in the nucleolytic cleavage reaction.

• 한국식품과학회지
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• 제25권1호
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• pp.46-51
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• 1993

본 연구는 2가철-아스코르빈산착체(Fe-Asc)에 의한 파아지 불활화에 있어서 Fe-Asc의 작용부위에 관해 연구하여 다음과 같은 결과를 얻었다. Fe-Asc의 단백질에 대한 작용에 관하여 우혈청알부민과 J1파아지의 구조단백질을 사용하여 검토한 결과 Fe-Asc의 처리구와 미처리구에 있어서 SDS-폴리아크릴아미드 겔 전기영동패턴, 아미노산 조성 및 자외선 스펙트럼에 변화는 보이지 않았다. 이에 대하여 Fe-Asc를 pUC18 DNA, M13mp8, ${\lambda}$ DNA 및 J1 파아지의 DNA에 작용시키면 아가로오스겔 전기영동패턴에 변화가 보여 사슬절단이 확인되었다. pUC18 DNA는 Fe-Asc와 반응시 먼저 수퍼코일형의 두가닥사슬 DNA의 한쪽 가닥에 절단이 일어나 개환형으로 되고 잇따라 두 가닥 사슬절단이 일어나 선형으로 되어 저분자화하는 것이 확인되었다. 이상의 결과로부터 Fe-Asc에 의한 파아지 불활화는 파아지 DNA의 손상에 기인한다고 생각된다.

• The Korean Journal of Physiology and Pharmacology
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• 제27권5호
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• pp.427-436
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• 2023

Mitotic arrest deficient 2 like 2 (Mad2L2, also known as Mad2B), the human homologue of the yeast Rev7 protein, is a regulatory subunit of DNA polymerase ζ that shares high sequence homology with Mad2, the mitotic checkpoint protein. Previously, we demonstrated the involvement of Mad2B in the cisplatin-induced DNA damage response. In this study, we extend our findings to show that Mad2B is recruited to sites of DNA damage in human cancer cells in response to cisplatin treatment. We found that in undamaged cells, Mad2B exists in a complex with Polζ-Rev1 and the APC/C subunit Cdc27. Following cisplatin-induced DNA damage, we observed an increase in the recruitment of Mad2B and Cdc20 (the activators of the APC/C), to the complex. The involvement of Mad2B-Cdc20-APC/C during DNA damage has not been reported before and suggests that the APC/C is activated following cisplatin-induced DNA damage. Using an in vitro ubiquitination assay, our data confirmed Mad2B-dependent activation of APC/C in cisplatin-treated cells. Mad2B may act as an accelerator for APC/C activation during DNA damage response. Our data strongly suggest a role for Mad2B-APC/C-Cdc20 in the ubiquitination of proteins involved in the DNA damage response.

• Journal of the Korean Society for Industrial and Applied Mathematics
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• 제15권3호
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• pp.161-175
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• 2011

An n-string tangle is a three dimensional ball with n strings properly embedded in it. A tangle model of a DNA-protein complex is first introduced by C. Ernst and D. Sumners in 1980's. They assumed the protein bound DNA as strings and the protein as a three dimensional ball. By using a tangle analysis, one can predict the topology of DNA within the complex. S.Kim and I. Darcy developed the biologically reasonable 4-string tangle equations and decided a solution tangle, called R-standard tangle. The author discussed more about the simple solution tangles of the equations and found a generalized R-standard tangle solution.

• 통합자연과학논문집
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• 제5권3호
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• pp.163-167
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• 2012

UV spectrophotometer was used to detect protein-DNA complex from DNA melting profile under constant temperature increase. Melting temperature (Tm) was $43^{\circ}C$ in copA duplex DNA alone. In the presence of Proteus mirabilis transcription regulator protein (PMTR) protein at 0.2 and 0.4 ${\mu}M$, Tm's were $45{\pm}0.5$ and $47.6{\pm}0.6^{\circ}C$, respectively. According to fluorescence polarization and gel shift assay. PMTR:copA complex was detected by the retarded migration on gel and the dissociation constant ($K_d$) was $(9.2{\pm}2.8){\times}10^{-9}M$.