The behaviour of dynein, a relatively little-studied protein found in muscle has been characterised using fluorescent markers and electron microscopy, paving the way for X-ray diffraction and NMR spectroscopy studies. The work not only shakes up the conventional wisdom on this motor protein but could shed new light on the debilitating symptoms and progression of motor neurone disease and related disorders.

Researchers in the University of Leeds' Astbury Centre for Structural Molecular Biology and at the University of Tokyo have, for the first time, identified key elements of dynein's structure, and visualised a "winch-like" mechanism by which it moves.

According to Leeds biologist Stan Burgess, dynein is the largest, but perhaps the least understood, of three families of motor proteins found in our bodies. Yet, despite the dearth of information on its properties and behaviour it plays a critical role in several processes. It provides the propulsion for the movement of sperm and eggs, for instance, and is involved in the process of cell division. The researchers point out that it is also involved in the transportation of chemicals within cells, including motor neurones, the nerve cells that supply all voluntary muscle activity, hence the direct connection with neurone degenerative diseases.

"Motor neurones have a very complex transportation system," explains Burgess, "While the nuclei of motor neurones lie within the spinal cord, they have branches that can run the entire length of a limb, say from the spine to the big toe. This branch is like a highway for molecular motors such as dynein. If there's a disruption to the traffic, it can lead to cell death and eventually to muscular weakness, characterised in diseases such as motor neurone disease."

Dynein, is a mere 50 nanometres in length but can carry its molecular cargo across distances of up to one metre in the human body. To give that distance a sense of scale, that would be the equivalent of a person walking approximately forty thousand kilometres. Until now, however, dynein structural studies have remained elusive because the protein engineering techniques required to make it amenable to analysis have not proved straightforward. Both X-ray crystallography and nuclear magnetic resonance spectroscopy have so far been unsuccessful.

The Leeds team has now worked with a synthetic dynein, the cytoplasmic dynein from Dictyostelium, successfully engineered by their colleagues in Japan. The synthetic protein contains fluorescent markers at key points within the molecular motor. These researchers could visualize these markers within the protein using electron microscopy. They were able to determine the positions in the presence and in the absence of adenosine triphosphate (ATP), the chemical "fuel" that drives the motor.

"Dynein, like all proteins, is a long linear molecule folded up into a complicated three-dimensional structure," says Burgess, "While we cannot solve the atomic structure using electron microscopy, our research has enabled us to map key points in the chain and see which parts of it move." According to team member Anthony Roberts, "Seeing the molecule change shape with ATP gives us clues to its motor mechanism that we will follow up in future work."

The scientists in Tokyo also removed the ends of the dynein molecule in one version of the engineered protein. This exposes the core allowing the Leeds researchers to get an insider view of the protein. In stark contrast to the conventional wisdom, the Leeds imaging of this exposed protein revealed the core to be similar to other ring-shaped molecular machines in the cell. Depending on the perspective taken, this is either a surprise or obvious-in-retrospect given that dynein shares distant evolutionary links with those other proteins.

"There has been disagreement over the structure of dynein within the scientific community, and both elements of our research - identifying the moving parts and revealing the structure of the core - has meant we can correct some of the mistaken ideas," explains Burgess, "Hopefully this will enable future research on this very important protein to move forward much faster."

The Leeds group now hopes to investigate the structure of two-headed dynein walking along its microtubule track using electron microscopy, while their colleagues in Tokyo measure the force the protein exerts during walking as well as its step size and speed. A new collaboration with a team in Ljubljana, Slovenia, will also allow them to combine the new data and simulate the movement of the protein.

"By examining the structure and mechanism of dynein while it's moving, we hope to learn more about how the protein works in the cell, so we can better understand what happens when it goes wrong," says Burgess.

Related links:

• Cell, 2009, 3, 485-495
• Astbury Centre

Article by David Bradley

The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.