博碩士論文 982402005 詳細資訊




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姓名 陳美廷(Mei-Ting Chen)  查詢紙本館藏   畢業系所 物理學系
論文名稱
(Real-Time Measurement of Vibrio alginolyticus Polar Flagellar Growth)
檔案 [Endnote RIS 格式]    [Bibtex 格式]    至系統瀏覽論文 (2019-2-1以後開放)
摘要(中) 細菌鞭毛是根生長於細菌體外的細長管子,此管子會形成螺旋推進器用以驅
動細菌運動。鞭毛的形成是一個自我組成的過程,鞭毛蛋白從管子的基部通過管
子中心被送到遠端,當它一到達頂端便會立即成為鞭毛的一部份,進而使整根鞭
毛長度增加。
為了研究這個奈米尺度的自我組成系統,我們開發了一種快速標記鞭毛的方
法,我們應用具有高空間和時間解析度的活體螢光影像直接觀測溶藻弧菌鞭毛的
生長。與先前的研究相比,我們的結論顯示溶藻弧菌的鞭毛成長速度與長度有高
度相關性:鞭毛長度短於1500 奈米,鞭毛以每分鐘50 奈米的速度等速成長,隨
著鞭毛長度增長,其成長速度會變慢,首先急劇下降接著趨於平緩,最後,當鞭
毛長度為7500 奈米,鞭毛的成長速度為每分鐘9 奈米。
我們建立了一個名為injection-diffusion 的模型來解釋我們的結果,由於鞭毛蛋白在管子內短距離的擴散速度很快,所以短鞭毛的鞭毛生長速度主要是由基部
鞭毛蛋白輸送進管內的速率所決定;當鞭毛長度增長,鞭毛蛋白的擴散變為有速
度限制的,部分的單體會塞在管內,阻礙新的鞭毛蛋白進入管內,鞭毛的生長速
度因而劇烈地減慢。
我們的結果提供了細菌鞭毛的動態生長過程一個新的觀點。
摘要(英) Bacterial flagella are thin external tubular filaments that form helical propellers to drive swimming in bacteria. Forming flagella is a self-assembly process when partially unfolded flagellum monomers, flagellins, are pumped out and transported through the central channel from its base to the distal end. The secretion apparatus contains six membrane proteins (FlhA, FlhB, FliO, FliP, FliQ and FliR) and three
soluble proteins (FliH, FliI, and FliJ) shareing the same structure with type III secretion system which is used solely for secretion. As flagellin reaches the growing
end of flagellar filament, it crystallizes immediately and becomes the new extending part of the filament. In order to study this nanometer-size self-assembled system, we have developed a fast flagellar filament labeling assay. We applied in vivo fluorescent imaging to monitor directly the growth of Vibrio alginolyticus polar flagella with high
spatial and temporal resolution. In contrast to previous reports, we revealed that the flagellar growth rate of Vibrio alginolyticus is highly length-dependent. When
flagellar length is shorter than 1500 nm, the flagellum growth is at a constant rate (50 nm/min). As the flagellum grows longer, the growth rate decays. It drops sharply at
first and tends to ease in the long flagella region. It decreases to 9 nm/min when flagellar length is 7500 nm. We built an injection-diffusion model to explain our results. When the flagellum is short, its growth rate is determined primarily by the loading speed at the filament base, since diffusion of flagellin across the short distance in the channel is fast. When the flagellum grows longer, diffusion of flagellin becomes rate-limiting. It causes some flagellar monomers jammed in the channel.
III
Such blockage slows down the loading of new flagellins into the channel. Therefore,the flagellar growth rate drops dramatically. Our results shed new light on the dynamic building process of this complex extracellular structure.
關鍵字(中) ★ 鞭毛
★ 溶藻弧菌
關鍵字(英) ★ flagella
★ sheath
★ vibrio alginolyticus
★ flagellin
論文目次 Abstract I
Acknowledgement IV
Content V
List of Figures VIII
Chapter1 Introduction 1
1-1 Overview to Bacteria 1
1-1-1 Classification of Bacteria 1
1-1-2 Bacterial Division 3
1-1-3 Patterns of Motility 4
1-2 Flagellar Motor 7
1-2-1 Components and Structure 8
1-2-2 Function 10
1-2-2-1 Power Source 10
1-2-2-2 Torque-generating Units 12
1-2-2-3 Switching 13
1-3 Flagellar Filaments 14
1-3-1 Atomic Structure 16
1-3-2 Flagellation 19
1-3-3 Type III Flagellar Export 20
1-4 Vibrio alginolyticus 22
1-5 Background and Models 24
Chapter2 Experimental Techniques 32
2-1 Apparatus 32
2-1-1 Atomic Force Microscope 32
2-1-2 Confocal Microscope 34
2-1-3 Epifluorescence Microscope 36
2-1-3-1 Inverted Optical Microscope 36
2-1-3-2 Fluorescence Microscope 38
2-1-3-3 Epifluorescence Microscope 39
2-2 materials and Methods 40
2-2-1 Strain Growth and Preparation 40
2-2-1-1 AFM Sample Preparation 40
2-2-1-2 Confocal Sample Preparation 40
2-2-1-3 Epifluorescence Sample Preparation 41
2-2-1-4 Buffer and Medium Contents 41
2-2-2 NanoOrange 42
2-2-3 Data Collection and Analysis 42
Chapter3 Experimental Results 44
3-1 AFM Morphology Study 44
3-1-1 Scanning in Air 44
3-1-2 Scanning in Aqueous Environment 45
3-2 NanoOrange Study 47
3-2-1 Staining on Bacteria 48
3-2-2 Binding Efficiency 48
3-2-3 Growing with NanoOrange 50
3-3 Population Study 52
3-4 Single Cell Study 55
3-4-1 Measuring Flagellum Length 56
3-4-2 Tethered Flagella 57
3-4-3 Non-tethered Flagella 59
3-4-4 Continuous Time-lapse Study 60
Chapter4 Discussion 62
4-1 Our Model 62
4-1-1 Constructing Model 62
4-1-2 Model Parameter Space Search 65
4-2 Flagellar Sheath 70
4-3 Energy Estimation for Pushing Flagellins into the Channel 72
4-4 Comparison with Background Researches 73
Chapter5 Conclusion 76
References 78
參考文獻 Aizawa SI and Kubori T (1998). "Bacterial flagellation and cell division." Genes Cells 3: 10.
Anderson RA and Berg HC (1973). "Bacteria swim by rotating their filaments." Nature 245: 3.
Bai F, Branch RW, Nicolau DV Jr, Pilizota T, Steel BC, Maini PK and Berry RM(2010). "Conformational spread as a mechanism for cooperativity in the bacterial flagellar switch." Science 327: 5.
Berg HC (2003). "The rotary motor of bacterial flagella." Annu Rev Biochem 72: 36.
Berg HC (2003). E. coli in Motion.
Blair DF (2003). "Flagellar movement driven by proton translocation." FEBS Letters 545: 10.
Blocker A, Komoriya K and Aizawa S (2003). "Type III secretion systems and bacterial flagella: Insights into their function from structural similarities." PNAS
100: 4.
Chen MT and Lo CJ (2016). Using biophysics to monitor the exxential protonmotive force in bacteria. Biophysics of Infection: 11.
Chen X and Berg HC (2000). "Torque-speed relationship of the flagllar rotary motor of Escherichia coli." B. J. 78: 6.
Diepold A and Armitage JP (2016). "Type III secretion systems: the bacterial flagellum and the injectisome." Phil. Trans. R. Soc. B 370: 19.
Erchardt M, Namba K, Hughes KT (2010). "Bacterial Nanomachines: Th flagellum and Type III injectisome." Cold Spring Harb Perspect Biol. 2:1
Evans LD, Poulter S, Terentjey EM, Hughes C and Fraser GM (2013). "A chain mechanism for flagellum growth." Nature 504: 16.
Francisi NR, Sosinsky GE, Thomas D and DeRosier DJ (1994). "Isolation,characterization and structure of bacterial flagellar motors containing the switch complex." J. Mol. Biol. 235: 10.
Furuno M, Atsumi T, Yamada T, Kojima S, Nishioka N, Kawagishi I and Homma M (1997). "Characterization of polar-flagellar-length mutants in Vibrio alginolyticus." Microbiology 143: 7.
Furuno M, Sato K, Kawagishi I and Homma M (2000). "Characterization of a flagellar sheath component, PF60 and its structural gene in marine vibrio." J.Biochem. 127(8): 29.
Galan JE (2008). "Energizing tpye III secretion machines: what is the fuel?" Nature Structure and Molecular Biology 15(2): 2.
Gosink KK and Hase CC (2000). "Requirements for conversion of the Na+-driven flagellar motor of vibrio cholerae to the H+-driven motor of Escherchia coli." J Bacteriol 182: 7.
Grossart HP, Steward GF, Martinez J and Azam F. "A simple, rapid method for demonstrating bacterial flagella." Appl. and Environment microbiol. 66(8):3632.
Harry E, Monahan L and Thompson L (2006). "Bacterial cell division: the mechanism and its precison." International Review of Cytology 253: 68.
Iino T (1974). "Assembly of Salmonella flagellin in vitro and in vivo." Jouranl of Supramolecular Structure 2: 13.
Jones CJ and Aizawa S (1991). "The bacterial flagellum and flagellar motor: structure, assembly and function." Advance in Microbial Physiol. 32: 7.
Kashket ER (1985). "The proton motive force in bacteria: a critical assessment of methods." Ann. Rev. Micro. 39: 24.
Keener JP (2006). "How Salmonella Typhimurium measures the length of flagellar filaments." Bull Math Biol 68: 18.
Khan S and Macnab RM (1980). "The steady-state counterclockwise/clockwise ratio of bacterial flagellar motors is regulated by protonmotive force." J. Mol. Biol.
138: 16.
Khan S, Dapice M and Reese TS (1988). "Effects of mot gene expression on the sturcture of the flagellar motor." J. Mol. Biol. 202: 10.
Kojima S, Kuroda M, Kawagishi I and Homma M (1999). "Random mutagenesis of the pomA gene encoding a putative channel component of the Na+-driven polar flagellar motor of Vibrio alginolyticus." Microbiology 145: 9.
Li N, Kojima S and Homma M (2011). "Sodium-driven motor of the polar flagellum in marine bacteria Vibrio." Genes Cells 16: 15.
Lin SN (2013). "長型群游菌的集體運動"
Macnab RM (2003). "How bacteria assemble flagella." Annu Rev Microbiol 57: 24.
Magariyama Y, Sugiyamas S, Muramoto K, Maekawa Y, Kawagishi I and Kudo S (1994). "Very fast flagellar rotation." Nature 371: 1.
McCarter L and Silverman M (1990). "Iron regulation of swarmer cell differentiation of Vibrioparahaemolyticus." J. Bacteriol. 171: 6.
McCarter L, Hilmen M and Silverman M (1988). "Flagellar dynamometer controls swarmer cell differentiation of V. parahaemolyticus." Cell 54: 7.
Meister M, Lowe G and Berg HC (1987). "The proton flux through the bacterial flagellar motor." Cell 49: 8.
Minamino T, Imada K and Namba K (2008). "Molecular motors of the bacterial flagella." Current Opinion in Structural Biology 18: 10.
Namba K, Yamashita I and Vonderviszt F (1989). "Structure of the core and central channel of bacterial flagella." Nature 342: 7.
Paul K, Brunstetter D, Titen S and Blair DF (2011). "A molecular mechanism of direction switching in the flagellar motor of Escherichia coli." PNAS 108: 6.
Paul K, Erhardt M, Hirano T, Blair DF and Hughes KT (2008). "Energy source of flagellar type III secretion." Nature 451: 5.
Reid SW, Leake MC, Chandler JH, Lo CJ, Armitage JP and Berry RM (2006). "The maximum number of torque-generating units in the flagellar motor of Escherichia coli is at least 11." PNAS 103: 6.
Sakar MK, Paul K and Blair D (2010) Chemotaxis signaling protein CheY binds to the rotor protein FliN to control the direction of flagellar rotation in Escherichia coli. PNAS 107:9370
Samatey FA, Imada K, Nagashima S, Vonderviszt F, Kumasaka T, Yamamoto M and Namba K (2001). "Structure of the bacterial flagellar protofilament and implications for a switch for supercoiling." Nature 410: 7.
Sjoblad RD, Emala CW and Doetsch RN (1983). "Invited review: bacterial flagellar sheaths: structures in search of a function." Cell Motility 3:93
Stern AS and Berg HC (2013). "Single-file diffusion of flagellin in flagellar filaments." Biophys J 105: 3.
Tanner DE, Ma W, Chen Z and Schulten K (2011). "Theoretical and computational investigation of flagellin translocation and bacterial flagellum growth." Biophys J 100: 9.
Tohru Minamino, K. I. a. K. N. (2008). "Mechanisms of type III protein export for bacterial flagellar assembly." Mol Biosyst 4: 11.
Turner L, Ryu WS and Berg HC (2000). "Real-time imaging of fluorescent flagellar filaments." J Bacteriol 182: 9.
Turner L, Stern AS and Berg HC (2012). "Growth of flagellar filaments of Escherichia coli is indenpendent of filament length." J Bacteriol 194: 6.
Weiss SJ and Weiss DS (2004). "Inhibiting cell division in Escherichia coli has little if any effect on gene expression." J Bacteriol 186: 5.
Yonekura K, Maki-Yonekura S and Namba K (2003). "Complete atomic model of the bacterial flagellar filament by electron cryomicroscopy." Nature 424: 8.
Yorimitsu T and Homma M (2001). "Na+-driven flagellar motor of Vibrio." BBA 1505: 12.
Zhu S, Kojima S and Homma M (2013). "Structure, gene regulation and environmental response of flagella in Vibrio." Front Microbiol 4: 9.
指導教授 羅健榮(Chien-Jung Lo) 審核日期 2017-1-23
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