Crystal structure of the hemochromatosis protein HFE...
Image: Laboratory of Pamela Bjorkman
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Hendrickson and his colleagues developed the MAD method in the 1980s and set up the Brookhaven beamline in part to do MAD experiments and see how they worked. "We knew the beamline would be useful anyway, even if the MAD idea turned out to have less promise than we hoped," says Hendrickson. "But we set it up especially so we could tune the x-ray wavelengths conveniently across the energy region. It was a slow process for us to get it together and we made a lot of mistakes along the way. But as it turned out, the Brookhaven beamline was extremely helpful to demonstrate to the world how valuable the technique was."
The first experiments were done in 1992. By 1993 Hendrickson and his colleagues were generating seminal papers. "It was fairly dramatic growth after that," he says. "And it's fair to say that the Hughes beamline is one of, if not the most productive of all the beamlines at Brookhaven—in large measure because a lot of people from Hughes have taken advantage of the capability."
Having seen how productive the Brookhaven beamline could be, Hughes investigators on the West Coast began lobbying for a more convenient facility. Protein crystals, after all, were still highly fragile and a cross-country trip to determine a structure could end in frustration. It was not uncommon, says HHMI investigator Pamela Bjorkman at the California Institute of Technology, for researchers to fly to the East Coast to use the Brookhaven beamline, only to find that their protein crystals, shipped in liquid nitrogen or taken at room temperature on the plane, had been lost by the shipping company or damaged en route.
"We had several experiences in which our crystals did not survive the travel," she recalls. "The entire trip would be a waste of money, time and crystals. We thought it would be nice to have a centrally located Hughes beamline or West Coast beamline. You could do a lot more things if you could routinely go to a synchrotron and not have to plan for a year in advance for your two days of beam time."
In 1998, HHMI investigators suggested to the Institute that it finance a beamline at the ALS. The ALS began operating in 1993 as a soft x-ray source for applications such as material science and x-ray microscopy. Although perfect for these techniques, soft x-rays are inappropriate for x-ray crystallography. From the outset, though, ALS engineers and physicists, working with a European colleague, were developing methods to produce higher-energy x-rays—hard x-rays in the physicists' vernacular—that are ideal for x-ray crystallography in general and MAD experiments in particular. The solution was to replace several of the magnets that bend electrons around the synchrotron ring with superconducting magnets, making it possible to provide more oomph at less expense in the same amount of space.
The West Coast investigators suggested that with the new, relatively low-cost technology, Hughes could fund a beamline that ALS personnel could run, unlike the Brookhaven beamline that is run and maintained by Hughes employees. The Institute's Trustees gave the go-ahead this year to begin funding two Hughes beamlines at the ALS at a cost of $8.05 million. The beamlines should be finished in about 18 months.
The new beamlines will join several dozen others that are in the works or planned at synchrotron sources like the ALS and others around the country. (more information)The new lines—and new techniques like MAD—will generate an explosion of structural data over the next decade, fundamentally altering our understanding of biology. "For Hughes investigators, the main application has been to look at increasingly more complicated biological systems," says Hendrickson.
The explosion of data and the speed at which it can be collected is also changing the way biologists work. Crystallographic studies used to be so ponderously slow that they lagged behind much of biological discovery. But as Kappler found out, once a structure was determined, the biology broke open. "Now we increasingly find scenarios where the crystallographic result is contemporaneous with or even leading the discovery phase," says Hendrickson. "The crystal structure gives you insight into the possible biology and then that directs experiments you do to understand what the biological action of this particular molecule might be."
"Even in situations where you already know a great deal about the biology of the system, the crystallographic insight gives you numerous ways of understanding the hows, the mechanisms by which the biochemistry underlying that biology is actually carried out," he adds. "All of this is having a profound impact."
