Douglas Rees is interested in the molecular mechanisms of bioenergetic processes, using x-ray crystallography and functional approaches. When he first started in protein x-ray crystallography in the mid-1970s, however, the future of this field didn't look bright.
“In some sense, the field appeared to be dying at that time,” he says. “There were a limited number of proteins naturally present in high abundance that you could study that restricted the appeal of subsequent crystal structure determinations. One reflection of this situation was that there were no academic jobs available to senior graduate students ahead of me.”
To use x-ray crystallography with proteins, researchers first need to obtain crystals, which historically has required large amounts of the protein of interest. Before genetic engineering, there was no way to selectively produce proteins in the lab, so researchers worked with the few naturally abundant proteins.
Rees almost avoided x-ray crystallography altogether. “I came to Harvard with a negative attitude toward crystallography, which was based on misconceptions,” he says. But a first-semester course on protein structure and function (taught by Stephen Harrison and the late Don C. Wiley, both of whom later became HHMI investigators) caught his attention.
“I learned about the protein-folding problem”—how proteins manage to fold themselves predictably and reliably into the proper conformations, seemingly without instructions—“and that seemed like about the most interesting problem there was,” he says. “I thought if you worked out the structures of the folded states, you could get some insights into this process.”
While a rotation student in Harrison’s lab, Rees discovered that he enjoyed structural research. “You could be involved in all sorts of activities—wet biochemistry, theory, instrumentation, computers—but you didn’t have to be an expert in every facet of the work. It’s more of a jack-of-all-trades sort of thing, which I happen to like.”
Following this rotation, Rees began his fascination with metalloproteins (proteins that contain metal ions) as a graduate student in the laboratory of William Lipscomb. Rees began analyzing the structures of inhibitors that bind to the active site of the zinc-containing enzyme carboxypeptidase. During this period, he was first exposed to the challenges of nitrogenase through extensive discussions with a sabbatical visitor to the Lipscomb lab, James B. Howard from the University of Minnesota. Nitrogenase is the only enzyme known to convert nitrogen gas to compounds such as nitrate and ammonia.
After postdoctoral stints with Lipscomb and Howard, Rees joined the faculty of the University of California, Los Angeles, in the Department of Chemistry and Biochemistry in 1982. In 1989, he moved to the California Institute of Technology, at a time when technological advances made it possible, at least in principle, to determine the structures of almost any protein. “At that time, almost anything you did was exciting since it hadn’t been done before.”
An important current focus of the Rees laboratory concerns the group of ABC (ATP binding cassette) transporter proteins, which are involved in all aspects of life. In bacterial systems, a major group of ABC transporters import nutrients into cells. Another group pumps molecules out of cells; related proteins in humans can lead to chemotherapy resistance in cancer patients. ABC transporters have also been implicated in genetic diseases. One important example involves mutations in a transporter gene called CFTR, which is responsible for cystic fibrosis.
ABC transporters move molecules across membranes using energy from ATP (adenosine triphosphate). How ABC transporters use ATP is not fully understood. Rees and colleagues have been studying bacterial transporters that function to import nutrients such as vitamin B12 and amino acids into cells. Qualitatively, the transport mechanism appears to resemble an airlock, with a series of “doors” that are opened and closed.
Despite challenges, Rees continues working to discover more about ABC transporters and other membrane proteins, including those from human sources. It’s difficult to make enough of the human proteins, he says, and there are also sometimes problems associated with detergents, which are used to extract the proteins from cell membranes. Rees is working with a colleague, Rob Phillips, and with Michael Stowell, a former student, and Hubert Yin at the University of Colorado, on an HHMI-funded Collaborative Innovation Award to develop methods for the structural analysis of membrane proteins in membrane bilayers that might be useful for both electron microscopy and x-ray crystallography studies.
Discovering more about these sentinels of the cell will provide crucial information that could help find new treatments for disease as well as advance our basic understanding of cells.
Though he finds his work fascinating and fulfilling, Rees did not plan his career as much as he took advantage of unexpected developments. “Much of what has happened in my life was not the consequence of a careful, long-range analysis of what I should be doing,” Rees says. “When an exciting and unanticipated opportunity came along, I was fortunate to be able to pursue it.”