Abstract
Bacterial flagella are extracellular filaments that drive swimming in bacteria. During motor assembly, flagellins are transported unfolded through the central channel in the flagellum to the growing tip. Here, we applied in vivo fluorescent imaging to monitor in real time the Vibrio alginolyticus polar flagella growth. The flagellar growth rate is found to be highly length-dependent. Initially, the flagellum grows at a constant rate (50 nm/min) when shorter than 1500 nm. The growth rate decays sharply when the flagellum grows longer, which decreases to ~9 nm/min at 7500 nm. We modeled flagellin transport inside the channel as a one-dimensional diffusive process with an injection force at its base. When the flagellum is short, its growth rate is determined by the loading speed at the base. Only when the flagellum grows longer does diffusion of flagellin become the rate-limiting step, dramatically reducing the growth rate. Our results shed new light on the dynamic building process of this complex extracellular structure.
Highlights
Protein transport in biology can involve active transport by molecular machines and passive transport through diffusion (Rapoport, 2007; Yuan et al, 2010; Jacobson et al, 1987)
Utilizing energy derived from ATP hydrolysis and proton motive force (PMF) (Paul et al, 2008; Minamino and Namba, 2008; Lee and Rietsch, 2015), the T3FSS continuously unfolds and delivers flagellin to the hollow interior of the motor, whereupon each unfolded flagellin protein is transported to the end of the filament and there
Accurate real-time measurement of flagellar growth in live cells is challenging because (1) flagella are thin extracellular filaments and there is not the contrast to observe the flagellar growth under bright-field microscopy
Summary
Protein transport in biology can involve active transport by molecular machines and passive transport through diffusion (Rapoport, 2007; Yuan et al, 2010; Jacobson et al, 1987). Using the transmembrane electrochemical proton motive force (PMF) to power the bacterial flagellar motor (BFM) (Manson et al, 1977; Gabel and Berg, 2003), fast rotating flagella can propel their cell body at a speed of 15–100 mm/s (Xue et al, 2015). Usually about 3–10 times the length of the cell body, are hollow protein cylinders of 20 nm outer diameter and 2 nm inner diameter (Yonekura et al, 2003). This long extracellular organelle is self-assembled from thousands of flagellin monomers (Figure 1a) (Yonekura et al, 2003; Asakura, 1970). Utilizing energy derived from ATP hydrolysis and PMF (Paul et al, 2008; Minamino and Namba, 2008; Lee and Rietsch, 2015), the T3FSS continuously unfolds and delivers flagellin to the hollow interior of the motor, whereupon each unfolded flagellin protein is transported to the end of the filament and there
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