Bacterial ﬂagella are extracellular ﬁlaments that drive swimming in bacteria. During its 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 (50nm/min) when shorter than 1500nm. The growth rate decays sharply when the ﬂagellum grows longer. 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.
- Chien-Jung Lo
- Fan Bai
- Fan Bai
- Chien-Jung Lo
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
- Richard M Berry, University of Oxford, United Kingdom
© 2017, Chen et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Mimivirus is the prototype of the Mimiviridae family of giant dsDNA viruses. Little is known about the organization of the 1.2 Mb genome inside the membrane-limited nucleoid filling the ~0.5 µm icosahedral capsids. Cryo-electron microscopy, cryo-electron tomography, and proteomics revealed that it is encased into a ~30-nm diameter helical protein shell surprisingly composed of two GMC-type oxidoreductases, which also form the glycosylated fibrils decorating the capsid. The genome is arranged in 5- or 6-start left-handed super-helices, with each DNA-strand lining the central channel. This luminal channel of the nucleoprotein fiber is wide enough to accommodate oxidative stress proteins and RNA polymerase subunits identified by proteomics. Such elegant supramolecular organization would represent a remarkable evolutionary strategy for packaging and protecting the genome, in a state ready for immediate transcription upon unwinding in the host cytoplasm. The parsimonious use of the same protein in two unrelated substructures of the virion is unexpected for a giant virus with thousand genes at its disposal.
Electron bifurcation is a fundamental energy conservation mechanism in nature in which two electrons from an intermediate-potential electron donor are split so that one is sent along a high-potential pathway to a high-potential acceptor and the other is sent along a low-potential pathway to a low-potential acceptor. This process allows endergonic reactions to be driven by exergonic ones and is an alternative, less recognized, mechanism of energy coupling to the well-known chemiosmotic principle. The electron-bifurcating [FeFe] hydrogenase from Thermotoga maritima (HydABC) requires both NADH and ferredoxin to reduce protons generating hydrogen. The mechanism of electron bifurcation in HydABC remains enigmatic in spite of intense research efforts over the last few years. Structural information may provide the basis for a better understanding of spectroscopic and functional information. Here, we present a 2.3 Å electron cryo-microscopy structure of HydABC. The structure shows a heterododecamer composed of two independent ‘halves’ each made of two strongly interacting HydABC heterotrimers connected via a [4Fe–4S] cluster. A central electron transfer pathway connects the active sites for NADH oxidation and for proton reduction. We identified two conformations of a flexible iron–sulfur cluster domain: a ‘closed bridge’ and an ‘open bridge’ conformation, where a Zn2+ site may act as a ‘hinge’ allowing domain movement. Based on these structural revelations, we propose a possible mechanism of electron bifurcation in HydABC where the flavin mononucleotide serves a dual role as both the electron bifurcation center and as the NAD+ reduction/NADH oxidation site.