Courtesy Julian Brown/Brandeis Photography
"We have your dog," read the cut-and-paste ransom note taped to Chris Miller's office door one day in the late 1990s. A biophysicist and Howard Hughes Medical Institute investigator at Brandeis University in Waltham, Mass., Miller regularly brought his West Highland terrier, Charlie, to the lab. "They even left a picture of the dog holding that day's newspaper," he laughs. Charlie was eventually returned, but only after Miller paid up, producing a stack of fill-in-the-blank, signed recommendation letters: "____ is the best person I've ever had in my lab. ..."
Miller is actually generous and efficient with recommendation letters. The ransom was a random joke, and a very good one, says Miller, who has been the target of countless pranks during his 30-odd years at the bench. His favorites, though, are the biological surprises that have essentially directed his science. "One of the best things about being a research scientist is that you are always subject to be the butt of nature's jokes," he says. "This is where your strongly held convictions are shown to be nonsense by nature herself. And it's an absolute delight, because that process of being jerked around makes you learn a lot of new things."
POTASSIUM TRUMPS CALCIUM
Among other things, nature's sense of humor drew Miller into his career-long exploration of the structure and function of membrane proteins. As a postdoc in Efraim Racker's laboratory at Cornell University, Miller set out to study the activity of the calcium ATPase that uses ATP to pump calcium against its gradient. He spent months designing a method for inserting the purified pumps into an artificial lipid bilayer, a simplified system that he thought would allow him to measure the currents generated by the movement of the charged calcium ions across the membrane. The pumps were derived from rabbit skeletal muscle, and if the photo that hangs outside Miller's lab is to be believed, the prep involved inserting a bunny feet-first into a Waring blender. Then again, you can't believe everything you see in Miller's lab.
Within minutes of adding the purified skeletal muscle proteins to the artificial membrane, Miller saw an electrical signal. But, it wasn't coming from a calcium pump, because the membrane preparation was responding to potassium, not calcium and ATP. "I said, 'What the hell is this?"' Miller had somehow managed to isolate a potassium channel. "It wasn't clear why potassium channels were there. They weren't supposed to be there," he says. But he decided to follow nature's lead and switch to studying the behavior of this channel in his minimalist membrane system.
Of course, not everyone agreed with Miller's assessment of the situation at the time. "When I published my first paper showing these channels, nobody believed it," says Miller. That's because studies of proteins reconstituted into artificial bilayers, he says, "had a bad rep on the street." Researchers had been using artificial membranes to look at ionophores, the small peptide-like molecules that spontaneously insert into the bilayer and form channels or carriers that transport ions. But they had yet to succeed with a full protein. All they'd seen were artifacts.
Miller was not perturbed by the skepticism. "It was a great gift," he says. "It gave me five years to work on this system essentially by myself without any competition. No one else wanted to touch it." He did, however, wonder why he kept finding channels that "weren't supposed to be there." So Miller decided to use his artificial system to fish for another protein that should be easy to find: the acetylcholine receptor, located at neuromuscular junctions. The receptor responds to acetylcholine's release from neurons by permitting the flow of sodium and potassium ions across the muscle cell membrane. For a rich source of receptors, Miller turned to the torpedo ray, an organism he says is "basically a swimming, purified acetylcholine receptor."
Again, in the first experiment, he detected a signal. This time it was "a chloride channel that wasn't supposed to be there," he says. "In the subsequent 15 years, I never once saw acetylcholine receptor currents," he notes. "I don't know why." Miller and other researchers eventually began to see "things that were supposed to be there," such as the sodium and potassium channels that were known to reside in nerve cell membranes. But he turned his full attention to investigating the less conventional channels: the potassium channel from rabbit muscle and the chloride channel that turned out to be part of a large family of so-called CLC chloride channels.
Early in his studies, Miller worked to "develop tools to probe and poke and feel around in channels in the dark to figure out what they must look like inside." In one series of experiments, he was able to surmise that the membrane-spanning portion of the potassium channel was shaped like an hourglass, with large cavernous vestibules at either end of a short, narrow passageway through which the ions squeeze. Miller's conjecture was confirmed when Roderick MacKinnon, his former postdoc now at Rockefeller University, published the X-ray structure of the potassium channel, work that earned MacKinnon a Nobel Prize in 2003. Similarly, Miller also deduced that the chloride channels are shaped like a double-barrel shotgun, another observation that was borne out by a crystal structure that MacKinnon's group later produced.
THAT'S NO CHANNEL
Miller's recent experiments led him to yet another stunning discovery: The chloride channel he'd been studying wasn't a channel at all; it was an active pump. After working for years on the CLC chloride channel from torpedo rays, Miller and his colleagues discovered that bacteria produce a similar protein. It was this bacterial protein that MacKinnon and his postdoc Raimund Dutzler used to determine their CLC channel structure.
Miller had great trouble reconstituting the bacterial channel in his artificial bilayer system, even though, he says, "The preparation was good, clean, beautiful. I mean, it crystallized, for God's sake." When he finally coaxed the proteins into his artificial membrane and characterized their electrical properties with his postdoc Alessio Accardi, Miller realized that their behavior was "not consistent with ions going through a protein according to a channel-like diffusion mechanism."
Instead, the protein was behaving like a transporter, exchanging chloride ions from one side of the membrane with protons on the other side. "That's something a channel will never do," says Miller, "because a channel's just a hole." But the observation suggested an experiment. If the protein were really an active transporter, it should be able to move chloride ions against their concentration gradient, another thing that channels can't do. "We did the test that day," says Miller, and they confirmed that the bacterial chloride channel is really a pump.
"A lot of people might have missed this," says Rob Blaustein of Tufts University Medical School, another of Miller's former post-docs. "But Chris fixated on it and realized that this protein has to be a transporter. It was an important discovery."
"Chris finds everything when he's looking for something else," says Joseph Mindell, a former postdoc who is now a biophysicist at the National Institutes of Health. "He has an astounding ability to smell when something he doesn't expect is actually meaningful, when a surprising result is something worth paying attention to and going after, and is not just an anomaly."
Of course, some members of the CLC family, including the protein from torpedo rays, are "true, kosher ion channels," says Miller. Others appear to be pumps, and Miller is now bent on determining why. He's using a directed approach to generate mutations that he hopes will transform the transporter into a channel. "If we can screw around here and screw around there and convert this thing into an ion channel, I think we'll actually understand something pretty important about how these proteins work," he says.
LETTING STRUCTURE DRIVE
After spending a couple of sabbaticals learning the basics of X-ray crystallography from MacKinnon and Dutzler, who's now at the University of Zurich, Miller is ready to tackle the structural questions head on. His colleagues are confident that he'll unearth something good. "Chris has an eye for breaking new ground," says Blaustein.
"He has a special skill for looking to the heart of a problem and asking questions that will let light into an area that had previously been dark," adds Steve Goldstein, a former postdoc who's now chair of pediatrics at the University of Chicago. "He does incredibly important work but he's not a salesman."
"Chris is a brilliant scientist. He's a gifted experimentalist and his science is superb," says Ronald Kaback, a friend and colleague at the University of California, Los Angeles. "He's one of my favorite people," adds Kaback, even though Miller stole the batteries from his laser pointer at a Keystone conference and convinced a fellow conferee to operate a substitute pointer surreptitiously from the audience. At first Miller's cocon-spirator directed the laser to the appropriate spot, so Kaback didn't immediately realize that his pointer had been disabled. But then the laser started to drift. "It slowly began to dawn on him that something wasn't right," says Miller.
"It was uproariously funny," admits Kaback, who once sent Miller a collection of 250 reprints by FedEx – C.O.D. That costly gag was a response to Miller's having, on a different occasion, sent Kaback a single reprint on which he had scrawled, "Free of charge." "You don't want science to get too serious," says Kaback.
Miller finds the research itself, particularly the electrophysiology, to be a blast. "It's like riding a horse. You see results in a few hundred milliseconds and you have to respond," he says. "Your nose is right in there with the channel and it feels like you're part of the experiment. It's still fun after all these years." And Miller is still active in the lab. "I got into this biz because I liked doing experiments," he says. "Why would I not do the thing I most like to do?"
For Miller, science is its own reward. He doesn't believe in special prizes, memberships, or accolades, says Kaback. Indeed, after attending the Nobel ceremony for his former postdoc MacKinnon, Miller says he "felt sorry for the poor schnooks who won." Their guests enjoyed a weeklong party, but the laureates were subjected to an endless series of formal events, giving presentations to the Swedish parliament and lunching with the queen. "It looked so painful," says Miller. He thinks all young scientists should be forced to attend. "It might dampen their Nobel lust," he says, and keep them doing science for the right reason: because it's fun.