BERKELEY, Calif. --In the short-term, the announcement is of consequence only to those cell biologists whose focus is on "mitochondria," the subcellular structures which generate energy for living cells. In the long-term, however, the announcement may be recorded as a milestone on the road to discovering a way to slow down the aging process, or a means of treating Alzheimer's disease or any of a number of other potentially devastating degenerative disorders.
An international team of researchers which includes scientists with the U.S. Department of Energy's Lawrence Berkeley National Laboratory has determined the complete crystal structure of one of the four protein complexes in the mitochondrial respiratory chain. Called cytochrome bc1 or complex III, this enzyme plays a critical role in the relay of electrons for producing energy that sustains the health of tissues, organs, and the body as a whole.
Bing Jap, a biophysicist with Berkeley Lab's Life Sciences Division, led the effort which took approximately eight years. Using x-ray crystallography, Jap and his colleagues have produced structural images of the entire 11 subunits of cytochrome bc1 at a resolution of approximately 3 angstroms. Images of the cytochrome bc1 complexes, which came from cow heart cells, provide the most detailed and complete pictures of the complex ever reported.
Every cell in the body contains hundreds of mitochondria which, together, provide about 90-percent of the energy that a cell needs to carry out its life processes. This energy is produced by the transfer of electrons from molecules of food through the respiratory chain and into the production of ATP, the molecules that served as traveling battery packs, delivering energy throughout the cell.
Numerous studies of the mitochondria have shown that anything that impedes the flow of electrons through the respiratory chain causes a decline in energy production. If uncorrected, this energy drop-off eventually begins to debilitate the normal operations of cells which, in turn, poses mounting problems for tissues and organs. Impeded electron flow also promotes the production of oxygen free-radical molecules which can attack all components of a cell and cause mutations in both nuclear and mitochondrial DNA.
As determined by Jap and his colleagues, the structure of cytochrome bc1 is that of a dimer, a complex of two neighboring molecular chains called monomers. The shape of the dimer is considered essential to its role in the electron-transfer process which is carried out in a hollow between its two monomers. This work has provided structural confirmation of the hollow's existence.
The cytochrome bc1 dimer is a non-crystalline protein embedded in the lipids of the mitochondrial inner membrane. Coaxing the protein into forming a crystal, a pre-requisite for x-ray imaging, was a major challenge.
"To crystallize cytochrome bc1 we had to first purify the protein by breaking the membrane using just the right proportion of a specific detergent," says Jap. "It is one of the most difficult of all crystallization techniques to carry out, but we now have one of the finest collections of (cytochrome bc1) crystals known. Without these high-quality crystals, we could not have achieved success."
With the solving of the cytochrome bc1 structure, scientists now have more than half of the mitochondrial respiratory chain filled in. What remains is the small complex II protein, and the very large complex I protein. Jap and his group will join the effort underway to solve complex I. For this, he plans to try a new strategy, one in which he will combine x-ray and electron crystallography techniques. This approach will include the use of multiple crystal averaging and phase information from both heavy atom derivatives and electron micrographs. Despite the small size of the complex II protein, its structure will also have to be solved.
As Jap explains, "Scientists need to have the resources to evaluate the entire structural picture of all the protein complexes in the mitochondrial respiratory chain in order to understand exactly what is going on. Otherwise, it is like trying to construct a map of Berkeley by visiting only half the city."
Collaborating with Jap on his work from Berkeley Lab were Joong Lee and John Kyongwon Lee, both with the Life Sciences Division. Other collaborators came from Sweden, Germany, and France. Their results were published in the July 3, 1998 issue of the journal Science (Vol. 281, pp. 64-71).
Berkeley Lab is a U.S. Department of Energy national laboratory located in Berkeley, California. It conducts unclassified scientific research and is managed by the University of California.