Assembly and operation mechanisms of flagellar apparatus of marine magnetotactic bacteria – MagneticFlAp

Bacterial flagella are nano-machines rotating at thousands-turns per second. Using genomic and biochemistry analyses we identify genes and proteins involved in its construction. Biophysical studies allow us to characterize sugars on the components of the flagella. Electron microscopy reveals detailed structure of this nano-machine. We analyze the swimming behavior in micro-fluidic systems under optical microscopes.

Our genomic analysis shows bacterial adaptations to marine habitats. In addition, collaboration between French and Japanese partners revealed an unprecedented complexity in protein composition and an exquisite architecture of the flagellar nano-machine of a Mediterranean bacterium. Seven flagella and 24 fibrils assemble into 7 intertwined hexagonal arrays within a sheath, which propels cells swimming at >100 body-length per second.

This research project will shed light into the operation mechanism of the flagellar nano-machine of marine magnetic bacteria. The results obtained have great potential of nano-technologic, biomedical and environmental applications.

The results obtained lead to three publications in peer-reviewed international journals in fields of molecular biology, environmental microbiology and general science (PNAS).

Bacteria sense a wide range of environmental signals and react to their changes by controlling their motility. The mostly used motility apparatus is the bacterial flagellum consisting of a basal body, a hook and a filament. The basal body contains a reversible rotary motor driven by either proton or sodium ion flow. The hook connects the motor to flagellar filament and plays a universal joint function. The flagellar filaments serve as a screw propeller to convert rotary motion of the motor into thrust. A flagellar filament is self-assembled from few tens of thousands copies of a single flagellin protein to achieve the helical structure by a mixture of the R- and L-type protofilaments. Each protofilament switches between these two conformations in response to a variety of environmental signals. The change of the signals affects thus the bacterial motility.
Magnetotactic bacteria (MTB) are capable of using the geomagnetic field as a cue for orientation in their searching for the optimal growth conditions. MTB consist of a heterogeneous group of gram-negative bacteria with various morphotypes. They synthesize peculiar prokaryotic organelles: magnetosomes. Magnetosomes comprise a magnetic single-domain crystal surrounded by a biomembrane and form chains inside cells. The chains impart a net magnetic dipole moment to the bacterium, allowing it to align and swim along geomagnetic field lines, a behavior called magnetotaxis. The proposed function of the magnetotaxis is that it increases the efficiency for bacteria to locate and maintain position at the oxic-anoxic transition zone (OATZ) of limnic or marine sediments. How flagella contribute to the magnetotaxis remains an open question.
Since the discovery of MTB in 1963, only a few axenic cultures are available because of their fastidious growth requirements. It severely hampers our understanding of magnetotaxis mechanism. We have successfully isolated a magneto-ovoid strain (MO-1), as the only axenic culture of MTB from the Mediterranean Sea. The MO-1 cells swim at an unusually high velocity (up to 300µm/sec), probably driven by both proton and sodium motors. There are two flagellar bundles on the same hemisphere of a cell. Each bundle consists of seven separated flagella that are enveloped by a common sheath. Our genomic analysis has identified 13 flagellin proteins. By using biochemistry analyses we have identified 3 glycosylated flagellins and 1 major glycosylated sheath protein. Little is known about protein glycosylation in prokaryotes, which is far behind our knowledge about eukaryotic cells. Prokaryotic glycoproteins have been found mainly in pathogens and play a vital role in pathogenicity and host invasion.
The structural and biochemical properties of the MO-1 flagellar apparatus require extremely exquisite system to transport the flagellins to the defined location and to precisely position the 7 flagella within the sheath. Moreover, protein secretion must be coordinated with the glycosylation process.
The MagneticFlAp project aims at elucidating the assembly and operation mechanisms of flagellar apparatus of MO-1 to shed light on the magnetotaxis mechanism. The ultimate challenge is to figure out how the MO-1 cells control the rotation of the flagella to respond to the change of the magnetic torque. It will be carried out by joining the three Partners having complementary expertise. Partner I is a well established and reference laboratory in the field of MTB. They have identified, for the first time, glycoproteins in MTB. Partner II has an internationally recognized expertise on the structural and functional analysis of complex carbohydrates. Partner III, Prof. Namba’s laboratory in Japan, is one of the best laboratories in the world working on bacterial flagellar apparatus. They have established the structural models of flagellar filament and hook of enterobacteria, which have been generally used as the reference structures.