Lorne Protein: Mr Namba's marvellous molecular machine

Graeme O'Neill learns about the ingenious engineering behind the molecular motor of the bacterial flagellum.

The Salmonella bacteria that cause food poisoning use propeller power to motor around in the digestive tracts of their unhappy hosts -- tiny, helical filaments of protein that generate thrust as they rotate.

The cells contain minuscule, rotary motors that project through the cell membrane, driving each of the microbe's multiple tails or flagellae, in a way that that permits them to rotate and flex through a wide range of orientations.

The system is a marvel of evolution, but it has also been held up by Creationists as a paragon of supernatural design, a prime exhibit in their case against Darwin's theory of evolution by natural selection.

Nature too moves in mysterious ways, but its designs are ultimately accessible to scientific scrutiny, even at the scale of the vanishingly small. Keiichi Namba, professor of the Graduate School of Frontier Bioscience at Osaka University in Japan, has probably done more than any other scientist to explain the genesis and workings of the marvellous molecular motor and its pliant propeller.

Hollywood

Namba's talk will be the highlight of the 30th annual Lorne Conference on Protein Structure and Function, according to conference publicity coordinator Dr Michael Parker, of St Vincent's Medical Research Institute in Melbourne. Parker said the dynamic Japanese researcher uses a Hollywood-class special-effects video to illustrate his talk describing how the motor and flagellum assemble themselves from protein components, and how the system works. The video is an animated synthesis of images of the structural elements of the nanomachine, obtained by X-ray crystallography and electron cryo-microscopy, combined with insights from computer-based simulations of its structure and workings.

Such is Namba's reputation that his peers named the research project after him -- he is director of the Namba Protonic NanoMachine Project, which is seeking to elucidate the structure and function of the bacterial flagellum as a huge molecular system, based on the organisation and movement of the individual atoms that build it. The project operates under the aegis of under Japan's ERATO (Exploratory Research for Advanced Technology) research program, established in 1981 to create new science and technologies for the Japanese economy, and stimulate interdisciplinary research and the development of new research tools and techniques.

ERATO projects tackle Big Questions. They provide a dynamic environment in which the cream of young, 30-plus Japanese PhDs and engineers can develop the broad knowledge and research and administrative skills required to excel on the world stage -- and Keiichi is a superstar graduate of the system. In October, Namba was senior author on a paper in Nature, 'Structure of the bacterial flagellar hook and implication for the molecular universal joint mechanism'. It describes his team's latest insights into the way the flexible 'tail' of the flagellum is coupled to the motor, and how it operates. The flagellum is assembled from about 25 different proteins, which form three components:

• A proton-powered rotary motor, assembled from a few to several tens of thousands of molecules, representing about 20 different proteins.
• The filament or 'tail', a long, tubular structure constructed from tens of thousands of copies of a single protein, flagellin (FliC), that coil and supercoil to form the flexible filament.
• A flagellar 'hook', linking the filament to the motor. The hook is a short, highly curved tubular structure, made from about 120 copies of a single protein, FlgE.

Two other proteins, called hook-associated proteins 1 and 3 (HAP1 and HAP3), connect the hook to the filament, which Namba and his colleagues believes acts as an adapter, providing a tight connection between the non-matching shapes and mechanical properties of the flexible hook and the relatively rigid filament.

Namba's team has shown that the hook linking the filament to the motor serves as a molecular universal joint -- yet another example of nature anticipating the ingenuity of human engineers, in this case by a few billion years. In their Nature paper, they noted that it essential for dynamic and efficient motility and taxis (movement towards food sources, or away from hostile environments).

The hook protein has a very different amino acid sequence to the filament protein, flagellin, but forms a very similar, tube-like helical structure. The authors comment, "It is curious that these two molecules with completely different structures both form tubular structures with basically the same architecture and helical symmetry".

Inherently flexible

The hook protein's unique 3D structure gives it inherently far more flexibility than flagellin, permitting it to "transmit torque from the motor to the helical propeller when the two are not coaxial". The flexibility allows the microbe to bundle its several filaments and rotate them synchronously in the same direction when it is 'swimming', or to rotate them independently, in different orientations, causing the microbe to exhibit a tumbling action.

The Japanese researchers used x-ray crystallography and electron cryo-microscopy to analyse the atomic structure of crystals of a large, core fragment of the 'hook' protein. They were limited to analysing the fragment, called FlgE31 because the intact protein cannot be crystallised -- it spontaneously forms filaments.

"The hook model now shows its complex molecular interactions and a possible switching mechanism to form a highly curved and twisted tubular structure within which individual protofilaments go through rather large and dynamic conformational changes for their rather extensive elongation and shortening as the curved hook rotates rapidly," they wrote.

It's not yet clear why the system requires not one but two 'adaptor' proteins, HAP1 and HAP2. Namba's team expects a structural analysis will reveal features matching the apposed regions of the hook protein and flagellin.

Namba will present the latest update in the tale of the marvellous microbial motor at the conference on Phillip Island. It should be of interest not just to protein researchers, but technologists looking for design ideas for self-propelled nanomachines.