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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 王惠鈞(Andrew H.-J. Wang) | |
dc.contributor.author | Chun-Han Ho | en |
dc.contributor.author | 何俊翰 | zh_TW |
dc.date.accessioned | 2021-06-16T02:54:03Z | - |
dc.date.available | 2015-07-20 | |
dc.date.copyright | 2015-07-20 | |
dc.date.issued | 2015 | |
dc.date.submitted | 2015-07-13 | |
dc.identifier.citation | Ali Azam, T., Iwata, A., Nishimura, A., Ueda, S., and Ishihama, A. (1999). Growth phase-dependent variation in protein composition of the Escherichia coli nucleoid. J Bacteriol 181, 6361-6370.
Ali, S.S., Beckett, E., Bae, S.J., and Navarre, W.W. (2011). The 5.5 protein of phage T7 inhibits H-NS through interactions with the central oligomerization domain. J Bacteriol 193, 4881-4892. Ali, S.S., Xia, B., Liu, J., and Navarre, W.W. (2012). Silencing of foreign DNA in bacteria. Curr Opin Microbiol 15, 175-181. Arold, S.T., Leonard, P.G., Parkinson, G.N., and Ladbury, J.E. (2010). H-NS forms a superhelical protein scaffold for DNA condensation. Proc Natl Acad Sci U S A 107, 15728-15732. Asensio, J.L., Perez-Lago, L., Lazaro, J.M., Gonzalez, C., Serrano-Heras, G., and Salas, M. (2011). Novel dimeric structure of phage phi29-encoded protein p56: insights into uracil-DNA glycosylase inhibition. Nucleic Acids Res 39, 9779-9788. Banos-Sanz, J.I., Mojardin, L., Sanz-Aparicio, J., Lazaro, J.M., Villar, L., Serrano-Heras, G., Gonzalez, B., and Salas, M. (2013). Crystal structure and functional insights into uracil-DNA glycosylase inhibition by phage Phi29 DNA mimic protein p56. Nucleic Acids Res 41, 6761-6773. Bhaya, D., Davison, M., and Barrangou, R. (2011). CRISPR-Cas systems in bacteria and archaea: versatile small RNAs for adaptive defense and regulation. Annu Rev Genet 45, 273-297. Bochkareva, E., Kaustov, L., Ayed, A., Yi, G.S., Lu, Y., Pineda-Lucena, A., Liao, J.C., Okorokov, A.L., Milner, J., Arrowsmith, C.H., et al. (2005). Single-stranded DNA mimicry in the p53 transactivation domain interaction with replication protein A. Proc Natl Acad Sci U S A 102, 15412-15417. Bouffartigues, E., Buckle, M., Badaut, C., Travers, A., and Rimsky, S. (2007). H-NS cooperative binding to high-affinity sites in a regulatory element results in transcriptional silencing. Nature structural & molecular biology 14, 441-448. Brunger, A.T., Adams, P.D., Clore, G.M., DeLano, W.L., Gros, P., Grosse-Kunstleve, R.W., Jiang, J.S., Kuszewski, J., Nilges, M., Pannu, N.S., et al. (1998). Crystallography & NMR system: A new software suite for macromolecular structure determination. Acta crystallographica Section D, Biological crystallography 54, 905-921. Castang, S., McManus, H.R., Turner, K.H., and Dove, S.L. (2008). H-NS family members function coordinately in an opportunistic pathogen. Proc Natl Acad Sci U S A 105, 18947-18952. Chen, C.C., Chou, M.Y., Huang, C.H., Majumder, A., and Wu, H.Y. (2005). A cis-spreading nucleoprotein filament is responsible for the gene silencing activity found in the promoter relay mechanism. J Biol Chem 280, 5101-5112. Comeau, S.R., Gatchell, D.W., Vajda, S., and Camacho, C.J. (2004a). ClusPro: a fully automated algorithm for protein-protein docking. Nucleic Acids Res 32, W96-99. Comeau, S.R., Gatchell, D.W., Vajda, S., and Camacho, C.J. (2004b). ClusPro: an automated docking and discrimination method for the prediction of protein complexes. Bioinformatics 20, 45-50. Court, R., Cook, N., Saikrishnan, K., and Wigley, D. (2007). The crystal structure of lambda-Gam protein suggests a model for RecBCD inhibition. J Mol Biol 371, 25-33. Cowtan, K. (2006). The Buccaneer software for automated model building. 1. Tracing protein chains. Acta crystallographica Section D, Biological crystallography 62, 1002-1011. Cowtan, K.D., and Main, P. (1996). Phase combination and cross validation in iterated density-modification calculations. Acta crystallographica Section D, Biological crystallography 52, 43-48. Dame, R.T., Wyman, C., and Goosen, N. (2000). H-NS mediated compaction of DNA visualised by atomic force microscopy. Nucleic Acids Res 28, 3504-3510. Dame, R.T., Wyman, C., and Goosen, N. (2001). Structural basis for preferential binding of H-NS to curved DNA. Biochimie 83, 231-234. Daubin, V., Lerat, E., and Perriere, G. (2003). The source of laterally transferred genes in bacterial genomes. Genome biology 4, R57. DeLano, W.L. (2008). The PyMOL Molecular Graphics System, Version 1.1. DeLano Scientific LLC, Palo Alto, CA, USA. Dharmalingam, K., Revel, H.R., and Goldberg, E.B. (1982). Physical mapping and cloning of bacteriophage T4 anti-restriction endonuclease gene. J Bacteriol 149, 694-699. Dryden, D.T., and Tock, M.R. (2006). DNA mimicry by proteins. Biochem Soc Trans 34, 317-319. Dutta, A., Ruppert, J.M., Aster, J.C., and Winchester, E. (1993). Inhibition of DNA replication factor RPA by p53. Nature 365, 79-82. Emsley, P., and Cowtan, K. (2004). Coot: model-building tools for molecular graphics. Acta crystallographica Section D, Biological crystallography 60, 2126-2132. Ericsson, U.B., Hallberg, B.M., Detitta, G.T., Dekker, N., and Nordlund, P. (2006). Thermofluor-based high-throughput stability optimization of proteins for structural studies. Anal Biochem 357, 289-298. Ghosh, M., Meiss, G., Pingoud, A.M., London, R.E., and Pedersen, L.C. (2007). The nuclease a-inhibitor complex is characterized by a novel metal ion bridge. J Biol Chem 282, 5682-5690. Gordon, B.R., Li, Y., Cote, A., Weirauch, M.T., Ding, P., Hughes, T.R., Navarre, W.W., Xia, B., and Liu, J. (2011). Structural basis for recognition of AT-rich DNA by unrelated xenogeneic silencing proteins. Proc Natl Acad Sci U S A 108, 10690-10695. Gordon, B.R., Li, Y., Wang, L., Sintsova, A., van Bakel, H., Tian, S., Navarre, W.W., Xia, B., and Liu, J. (2010). Lsr2 is a nucleoid-associated protein that targets AT-rich sequences and virulence genes in Mycobacterium tuberculosis. Proc Natl Acad Sci U S A 107, 5154-5159. Grove, A. (2011). Functional evolution of bacterial histone-like HU proteins. Curr Issues Mol Biol 13, 1-12. Hegde, S.S., Vetting, M.W., Roderick, S.L., Mitchenall, L.A., Maxwell, A., Takiff, H.E., and Blanchard, J.S. (2005). A fluoroquinolone resistance protein from Mycobacterium tuberculosis that mimics DNA. Science 308, 1480-1483. Kahramanoglou, C., Seshasayee, A.S., Prieto, A.I., Ibberson, D., Schmidt, S., Zimmermann, J., Benes, V., Fraser, G.M., and Luscombe, N.M. (2011). Direct and indirect effects of H-NS and Fis on global gene expression control in Escherichia coli. Nucleic Acids Res 39, 2073-2091. Kennaway, C.K., Obarska-Kosinska, A., White, J.H., Tuszynska, I., Cooper, L.P., Bujnicki, J.M., Trinick, J., and Dryden, D.T. (2009). The structure of M.EcoKI Type I DNA methyltransferase with a DNA mimic antirestriction protein. Nucleic Acids Res 37, 762-770. Kim, B.C., Kim, K., Park, E.H., and Lim, C.J. (1997). Nucleotide sequence and revised map location of the arn gene from bacteriophage T4. Mol Cells 7, 694-696. Kozakov, D., Brenke, R., Comeau, S.R., and Vajda, S. (2006). PIPER: an FFT-based protein docking program with pairwise potentials. Proteins 65, 392-406. Kozakov, D., Hall, D.R., Beglov, D., Brenke, R., Comeau, S.R., Shen, Y., Li, K., Zheng, J., Vakili, P., Paschalidis, I., et al. (2010). Achieving reliability and high accuracy in automated protein docking: ClusPro, PIPER, SDU, and stability analysis in CAPRI rounds 13-19. Proteins 78, 3124-3130. Lang, B., Blot, N., Bouffartigues, E., Buckle, M., Geertz, M., Gualerzi, C.O., Mavathur, R., Muskhelishvili, G., Pon, C.L., Rimsky, S., et al. (2007). High-affinity DNA binding sites for H-NS provide a molecular basis for selective silencing within proteobacterial genomes. Nucleic Acids Res 35, 6330-6337. Lawrence, J.G., and Ochman, H. (1997). Amelioration of bacterial genomes: rates of change and exchange. J Mol Evol 44, 383-397. Leon, E., Navarro-Aviles, G., Santiveri, C.M., Flores-Flores, C., Rico, M., Gonzalez, C., Murillo, F.J., Elias-Arnanz, M., Jimenez, M.A., and Padmanabhan, S. (2010). A bacterial antirepressor with SH3 domain topology mimics operator DNA in sequestering the repressor DNA recognition helix. Nucleic Acids Res 38, 5226-5241. Liu, D., Ishima, R., Tong, K.I., Bagby, S., Kokubo, T., Muhandiram, D.R., Kay, L.E., Nakatani, Y., and Ikura, M. (1998). Solution structure of a TBP-TAF(II)230 complex: protein mimicry of the minor groove surface of the TATA box unwound by TBP. Cell 94, 573-583. Liu, Y., Chen, H., Kenney, L.J., and Yan, J. (2010). A divalent switch drives H-NS/DNA-binding conformations between stiffening and bridging modes. Genes Dev 24, 339-344. McMahon, S.A., Roberts, G.A., Johnson, K.A., Cooper, L.P., Liu, H., White, J.H., Carter, L.G., Sanghvi, B., Oke, M., Walkinshaw, M.D., et al. (2009). Extensive DNA mimicry by the ArdA anti-restriction protein and its role in the spread of antibiotic resistance. Nucleic Acids Res 37, 4887-4897. Navarre, W.W., McClelland, M., Libby, S.J., and Fang, F.C. (2007). Silencing of xenogeneic DNA by H-NS-facilitation of lateral gene transfer in bacteria by a defense system that recognizes foreign DNA. Genes Dev 21, 1456-1471. Navarre, W.W., Porwollik, S., Wang, Y., McClelland, M., Rosen, H., Libby, S.J., and Fang, F.C. (2006). Selective silencing of foreign DNA with low GC content by the H-NS protein in Salmonella. Science 313, 236-238. Niesen, F.H., Berglund, H., and Vedadi, M. (2007). The use of differential scanning fluorimetry to detect ligand interactions that promote protein stability. Nature protocols 2, 2212-2221. Ochman, H., Lawrence, J.G., and Groisman, E.A. (2000). Lateral gene transfer and the nature of bacterial innovation. Nature 405, 299-304. Otwinowski, Z.a.M., W. (1997). Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol 276. Parsons, L.M., Yeh, D.C., and Orban, J. (2004). Solution structure of the highly acidic protein HI1450 from Haemophilus influenzae, a putative double-stranded DNA mimic. Proteins 54, 375-383. Perkins, D.N., Pappin, D.J., Creasy, D.M., and Cottrell, J.S. (1999). Probability-based protein identification by searching sequence databases using mass spectrometry data. Electrophoresis 20, 3551-3567. Potterton, E., Briggs, P., Turkenburg, M., and Dodson, E. (2003). A graphical user interface to the CCP4 program suite. Acta crystallographica Section D, Biological crystallography 59, 1131-1137. Powell, L.M., Dryden, D.T., and Murray, N.E. (1998). Sequence-specific DNA binding by EcoKI, a type IA DNA restriction enzyme. J Mol Biol 283, 963-976. Putnam, C.D., Shroyer, M.J., Lundquist, A.J., Mol, C.D., Arvai, A.S., Mosbaugh, D.W., and Tainer, J.A. (1999). Protein mimicry of DNA from crystal structures of the uracil-DNA glycosylase inhibitor protein and its complex with Escherichia coli uracil-DNA glycosylase. J Mol Biol 287, 331-346. Putnam, C.D., and Tainer, J.A. (2005). Protein mimicry of DNA and pathway regulation. DNA repair 4, 1410-1420. Rajagopalan, S., Andreeva, A., Teufel, D.P., Freund, S.M., and Fersht, A.R. (2009). Interaction between the transactivation domain of p53 and PC4 exemplifies acidic activation domains as single-stranded DNA mimics. J Biol Chem 284, 21728-21737. Ramirez, B.E., Voloshin, O.N., Camerini-Otero, R.D., and Bax, A. (2000). Solution structure of DinI provides insight into its mode of RecA inactivation. Protein Sci 9, 2161-2169. Rimsky, S., Zuber, F., Buckle, M., and Buc, H. (2001). A molecular mechanism for the repression of transcription by the H-NS protein. Mol Microbiol 42, 1311-1323. Sheldrick, G.M. (2008). A short history of SHELX. Acta crystallographica Section A, Foundations of crystallography 64, 112-122. Smits, W.K., and Grossman, A.D. (2010). The transcriptional regulator Rok binds A+T-rich DNA and is involved in repression of a mobile genetic element in Bacillus subtilis. PLoS genetics 6, e1001207. Sommer, N., Salniene, V., Gineikiene, E., Nivinskas, R., and Ruger, W. (2000). T4 early promoter strength probed in vivo with unribosylated and ADP-ribosylated Escherichia coli RNA polymerase: a mutation analysis. Microbiology 146 ( Pt 10), 2643-2653. Sukackaite, R., Grazulis, S., Tamulaitis, G., and Siksnys, V. (2012). The recognition domain of the methyl-specific endonuclease McrBC flips out 5-methylcytosine. Nucleic Acids Res 40, 7552-7562. Swinger, K.K., Lemberg, K.M., Zhang, Y., and Rice, P.A. (2003). Flexible DNA bending in HU-DNA cocrystal structures. EMBO J 22, 3749-3760. Tendeng, C., Soutourina, O.A., Danchin, A., and Bertin, P.N. (2003). MvaT proteins in Pseudomonas spp.: a novel class of H-NS-like proteins. Microbiology 149, 3047-3050. Thresher, R., and Griffith, J. (1992). Electron microscopic visualization of DNA and DNA-protein complexes as adjunct to biochemical studies. Methods Enzymol 211, 481-490. Travers, A.A., Muskhelishvili, G., and Thompson, J.M. (2012). DNA information: from digital code to analogue structure. Philos Trans A Math Phys Eng Sci 370, 2960-2986. Tucker, A.T., Bobay, B.G., Banse, A.V., Olson, A.L., Soderblom, E.J., Moseley, M.A., Thompson, R.J., Varney, K.M., Losick, R., and Cavanagh, J. (2014). A DNA mimic: the structure and mechanism of action for the anti-repressor protein AbbA. J Mol Biol 426, 1911-1924. Tupper, A.E., Owen-Hughes, T.A., Ussery, D.W., Santos, D.S., Ferguson, D.J., Sidebotham, J.M., Hinton, J.C., and Higgins, C.F. (1994). The chromatin-associated protein H-NS alters DNA topology in vitro. EMBO J 13, 258-268. Vagin, A.A., Steiner, R.A., Lebedev, A.A., Potterton, L., McNicholas, S., Long, F., and Murshudov, G.N. (2004). REFMAC5 dictionary: organization of prior chemical knowledge and guidelines for its use. Acta crystallographica Section D, Biological crystallography 60, 2184-2195. Walkinshaw, M.D., Taylor, P., Sturrock, S.S., Atanasiu, C., Berge, T., Henderson, R.M., Edwardson, J.M., and Dryden, D.T. (2002). Structure of Ocr from bacteriophage T7, a protein that mimics B-form DNA. Mol Cell 9, 187-194. Wang, H.C., Ho, C.H., Hsu, K.C., Yang, J.M., and Wang, A.H. (2014a). DNA mimic proteins: functions, structures, and bioinformatic analysis. Biochemistry (Mosc) 53, 2865-2874. Wang, H.C., Hsu, K.C., Yang, J.M., Wu, M.L., Ko, T.P., Lin, S.R., and Wang, A.H. (2014b). Staphylococcus aureus protein SAUGI acts as a uracil-DNA glycosylase inhibitor. Nucleic Acids Res 42, 1354-1364. Wang, H.C., Ko, T.P., Lee, Y.M., Leu, J.H., Ho, C.H., Huang, W.P., Lo, C.F., and Wang, A.H. (2008). White spot syndrome virus protein ICP11: A histone-binding DNA mimic that disrupts nucleosome assembly. Proc Natl Acad Sci U S A 105, 20758-20763. Wang, H.C., Ko, T.P., Wu, M.L., Ku, S.C., Wu, H.J., and Wang, A.H. (2012). Neisseria conserved protein DMP19 is a DNA mimic protein that prevents DNA binding to a hypothetical nitrogen-response transcription factor. Nucleic Acids Res 40, 5718-5730. Wang, H.C., Wu, M.L., Ko, T.P., and Wang, A.H. (2013). Neisseria conserved hypothetical protein DMP12 is a DNA mimic that binds to histone-like HU protein. Nucleic Acids Res 41, 5127-5138. Werner, R.M., Jiang, Y.L., Gordley, R.G., Jagadeesh, G.J., Ladner, J.E., Xiao, G., Tordova, M., Gilliland, G.L., and Stivers, J.T. (2000). Stressing-out DNA? The contribution of serine-phosphodiester interactions in catalysis by uracil DNA glycosylase. Biochemistry (Mosc) 39, 12585-12594. Wilson, G.G., and Murray, N.E. (1991). Restriction and modification systems. Annu Rev Genet 25, 585-627. Zuber, S., Ngom-Bru, C., Barretto, C., Bruttin, A., Brussow, H., and Denou, E. (2007). Genome analysis of phage JS98 defines a fourth major subgroup of T4-like phages in Escherichia coli. J Bacteriol 189, 8206-8214. | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/54388 | - |
dc.description.abstract | DNA是一般生物的遺傳物質,經由一些DNA相關之功能性蛋白質,如轉錄因子或是DNA結合蛋白質可以調控DNA序列中所蘊涵的遺傳訊息之表達。大約在十年前,有一類的蛋白質稱之DNA擬態蛋白質被發現,他們在蛋白質結構上所展現出的特殊形態是模擬DNA的總體形狀以及DNA表面負電荷排列狀態,使其他與DNA作用的蛋白質無法區別而與之作用,進而達到抑制某些特定DNA相關功能性蛋白質的活性與功能,因此DNA擬態蛋白質往往參與著很多重要的細胞生理調控以及存在於許多尚未被發現的調控機制當中。
為了找尋T4噬菌體中DNA擬態蛋白質,在本篇研究中,先鎖定具有DNA擬態蛋白質的通性:分子量較小與等電點較低的蛋白質做為我們初篩選的標的蛋白質進行分析。經篩選後發現,其中Arn這個蛋白質很有可能是一個尚未被發現的DNA擬態蛋白質;於是利用結構生物學的方法,解析Arn的3D立體結構。非常有趣並令人振奮地,從Arn的3D立體結構可以發現:Arn二聚體表面帶負電的胺基酸分佈與雙股DNA帶負電的磷酸基團分佈極為類似,此結果強烈暗示著Arn可能是一個DNA擬態蛋白質。除此之外,Arn二聚體的大小與形狀也與另一個已知的DNA擬態蛋白質Ocr之二聚體相類似。為了解T4噬菌體Arn在大腸桿菌體中與哪個蛋白質有交互作用,於是進行了蛋白質沉澱技術,並發現細菌類組蛋白質H-NS可與T4噬菌體蛋白質Arn進行蛋白質-蛋白質專一性結合。 而這個初步的發現暗示著可能隱藏著全新的調控機制。由於T4噬菌體感染其染宿主大腸桿菌時會遭受到宿主的防禦系統攻擊,如:細菌體內會表現一些小型的DNA結合蛋白質結合外來或噬菌體DNA並造成抑制其基因表現,其中細菌類組蛋白質H-NS就是此類蛋白質的成員之一。因此T4噬菌體為了能成功地感其染宿主並在菌體內快速完成感染週期,T4噬菌體勢必要演化出抵抗宿主此類的防禦系統。而T4噬菌體利用DNA擬態蛋白質來抑制細菌類組蛋白質H-NS與噬菌體DNA結合,對噬菌體來說,則很可能是一個很有效率抵抗宿主的機制。 因此為了更進一步證實與了解T4噬菌體蛋白質Arn的功能是否與我們的假設相符,利用競爭性的電泳遷移率改變分析法,可以發現Arn可以競爭並取代DNA片段與DNA結合蛋白質H-NS結合;在電腦3D結構的模擬中更顯示著Arn二聚體利用模擬DNA電性分佈的一側和H-NS原本與DNA結合區域進行結合,進而阻擋了DNA與H-NS的結合。此外;在胞外基因表現分析與電子顯微鏡影像分析中更證實Arn可以阻擋H-NS對基因抑制的活性。 在此研究中,我們在T4噬菌體中發現了一個全新的噬菌體-宿主的對抗機制:一個全新的DNA擬態蛋白質Arn,它藉由模擬DNA表面負電荷分佈,與細菌類組蛋白質H-NS結合並阻擋了其DNA結合位,而達到阻擋H-NS對基因抑制的活性。 | zh_TW |
dc.description.abstract | Organisms using DNA as their hereditary substance, the genetic information of these DNA sequence can be expressed and regulated by DNA-related functional proteins, such as transcriptional factors and DNA-binding proteins. Around a decade ago, a new category of control factors of DNA-related functional proteins called the DNA mimic protein had been identified. The DNA mimic protein can inhibit and/or regulate the function and activity of DNA-related functional proteins by its DNA-like shape and unique surface negative charge pattern. Thus, DNA mimic proteins are involved in certain important cellular processes and may be involved in many undiscovered regulation mechanisms.
To identify the DNA mimic protein in T4 phage, we focused on the proteins which have the properties found in known DNA mimic proteins: small protein size and low isoelectric point (pI). After the sequence analysis, we found that anti restriction nuclease (Arn) protein may be a DNA mimic protein. By using structural approaches, the crystal structure of Arn was determined. Interestingly, the negative charge distribution of Arn dimer surface is similar to the phosphate group distribution on DNA, implying Arn dimer could act as a DNA mimic. Furthermore, the size and shape of Arn dimer is similar to the DNA mimic protein overcome classical restriction (Ocr). To identify the interaction partner of T4 phage Arn in Escherichia coli (E. coli), the His-pull-down was used and we further discovered that bacterial histone-like nucleoid structuring (H-NS) protein can interact with Arn specifically. Arn/H-NS interaction reveals a novel regulation mechanism. When infecting the E. coli, T4 phage encounters attacks from host’s defense systems. For example, bacteria express some small DNA-binding proteins to bind and entangle foreign or phage DNAs, inducing the gene silencing effect. H-NS is a member of these kind of protein. Thus, in order to infect and replicate in E. coli successfully, T4 phage has to evolve some strategies to overcome the gene silencing effect from H-NS. Therefore, using a DNA mimic protein to inhibit H-NS is a straightforward way for T4 phage. To confirm our hypothesis in this undiscovered function of Arn, electrophoresis mobility shift assay (EMSA) was used and demonstrated that Arn competes with bacteria as well as phage DNA fragments for binding to H-NS. Computer modeling analysis revealed that Arn dimer competes with DNA to interact with the H-NS DNA binding domain via its negatively-charged side. Additionally, in vitro gene expression and electron microscopy analyses further indicated that Arn antagonizes the gene-silencing effect of H-NS on the reporter gene. In summary, we discovered a novel mechanism for phage-bacteria battle from T4 phage, which employs the DNA mimic protein Arn to counteract the H-NS gene-silencing effect by its DNA-like surface. | en |
dc.description.provenance | Made available in DSpace on 2021-06-16T02:54:03Z (GMT). No. of bitstreams: 1 ntu-104-F97b46017-1.pdf: 20041130 bytes, checksum: 24256dcd636d47ff1c04b8e43bdc110a (MD5) Previous issue date: 2015 | en |
dc.description.tableofcontents | List of Tables……………………………………………………………….i
List of Figures…………………………………………………..…………ii Abbreviations………………………………………………..…………iv 中文摘要…………………………………………………………………...1 Abstract……………………………………………………………………3 Introduction 1-1 DNA mimic proteins…………………………………………………………………6 1-1-1 Long double-stranded DNA (dsDNA) mimic proteins………………….……8 1-1-2 Short double-stranded DNA mimic proteins………………………….………8 1-1-3 Single-stranded DNA (ssDNA) mimic proteins………..…………….………9 1-1-4 Other DNA mimic proteins..…………………………………..…….………10 1-2 Newly discovered T4 DNA mimic protein: Arn (anti restriction nuclease)...…….…12 1-3 Defense against exogenous DNAs and the gene silencer H-NS in bacteria……….13 Materials and methods 2-1 Preparation of recombinant Arn-His and H-NS………………...…………………..15 2-2 The thermal shift assay of Arn-His and His-Arn……………………………………16 2-3 Recombinant Arn-His crystallization and data collection…………………………..18 2-4 Structure determination and refinement…………………………………………….19 2-5 His-pull-down assay and protein identification……………………………………..19 2-6 Analytical ultracentrifugation (AUC) analysis……………………………………..21 2-7 Electrophoresis mobility shift assay (EMSA)………………………………………21 2-8 In vitro gene regulation assay……………………………………………………….22 2-9 Transmission electron microscopy (TEM) imaging……………………………...…23 2-10 Protein data bank accession code……………………………………………..….23 Results and discussion 3-1 Exploring the potential DNA mimic protein of T4 phage………………………..….24 3-2 The crystal structure of Arn shows DNA mimic properties…………………...…….24 3-3 E. coli H-NS interacts with Arn………………………………………...……...……26 3-4 Arn/H-NS binding is affected by salt concentration and/or pH that similar to the properties of DNA/H-NS interaction……………………………………………..…….28 3-5 Arn disrupts the binding between H-NS and DNA………………..………..……….29 3-6 The Arn protein neutralizes the gene-silencing effect of the H-NS protein………....31 3-7 The Arn protein affects the higher order structure of plasmid DNA induced by H-NS………………………………………………………………………....……………32 3-8 A proposed binding model shows how Arn interacts with the DNA binding domain of H-NS………………………………………………………………………...……….33 3-9 Arn mimics DNA substrate exquisitely………………..……………………...…….34 Conclusion………………………………………………………………..37 References………………………………………………………………..39 Tables…………………………………………………………..…………52 Figures……………………………………………………………………59 Appendices…………………………………………………………….…91 | |
dc.language.iso | en | |
dc.title | T4噬菌體DNA擬態蛋白質Arn抑制細菌類組蛋白質H-NS之DNA結合活性研究 | zh_TW |
dc.title | The T4 Phage DNA Mimic Protein Arn Inhibits the DNA-binding Activity of the Bacterial Histone-like Protein H-NS | en |
dc.type | Thesis | |
dc.date.schoolyear | 103-2 | |
dc.description.degree | 博士 | |
dc.contributor.oralexamcommittee | 詹迺立(Nei-Li Chan),張崇毅(Chung-I Chang),何孟樵(Meng-Chiao Ho),王皓青(Hao-Ching Wang) | |
dc.subject.keyword | 分子擬態,DNA擬態蛋白質,抗細菌(宿主)防禦機制,抗基因沉默,類核體, | zh_TW |
dc.subject.keyword | molecular mimicry,DNA mimic protein,anti-bacterial defense system,anti-gene silencing,nucleoid, | en |
dc.relation.page | 90 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2015-07-13 | |
dc.contributor.author-college | 生命科學院 | zh_TW |
dc.contributor.author-dept | 生化科學研究所 | zh_TW |
顯示於系所單位: | 生化科學研究所 |
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