RACHELLE GAUDET IS HOOKED ON CRYSTALLOGRAPHY
July 27th, 2006
But following Wiley’s untimely death in 2001 and the dissolution of his lab, Gaudet didn’t want the TAP project to die. So she adopted TAP as her own when she began her tenure as Assistant Professor in MCB in 2002. Now, four years later and recently promoted to Associate Professor, Gaudet has made gratifying progress on both her TAP and TRP projects, which both entail understanding how the structure of proteins affects their biological function. She combines that structural information with cellular biology and biochemistry to learn about the protein’s mechanism by studying mammalian proteins produced in cell cultures.
"I wanted to be an architect growing up until I became interested in science," Gaudet recalls. "In structural biology, we study the architecture of the protein. Nature tells you the architecture. You just have to display it properly."
Proteins start out as linear polypeptides that fold into three-dimensional shapes, Gaudet explains, and structural biology looks at how proteins do that folding. To reveal structure, she uses X-ray crystallography to discover exactly where every atom in a complex protein – or part of a protein – is arranged three-dimensionally.
Gaudet first became "hooked on crystallography" as an undergraduate at Université de Montréal when she realized you could look at a protein’s structure and figure out its function. "I finally knew how things worked at the level of detail I needed," she says. The most exciting moment was when she determined the structure of a new protein (phosducin, a protein in photoreceptors) as a PhD candidate in Molecular Biophysics and Biochemistry at Yale University. "It was a great thrill to look at the protein and know I was the first person to see it."
Around that time, Gaudet was discovering another thrill, a type of swing dancing called the Lindy Hop. An injury had kept her off the competitive swim team in college, so she found another outlet for that drive in dancing. When she arrived at Harvard, she continued studying dance at a local studio, Hop to the Beat, began competing and accumulating prizes, and later joined the T Token Swing dance team.
A Two-Cylinder Transporter
Gaudet’s interest in crystallography drew her to Wiley’s lab, where he and Jack Strominger, also of MCB, were involved in a long collaboration on major histocompatibility complexes (MHCs). They were working on how MHCs present peptides from viruses, microorganisms, or foreign or deregulated cells at the cell surface to alert immune cells about imminent danger. "My project went one step back in the pathway to discover how peptides are loaded on the MHC molecules," Gaudet says. She focused on TAP, which stands for transporter associated with antigen processing.
As a transporter, TAP creates a channel through the endoplasmic reticulum (ER) membrane that ushers peptides into the ER lumen and delivers them to the MHCs. Gaudet decided to first tackle just the intracellular domain of this large and complicated protein. This section houses a "two-cylinder engine" that drives the transport of peptides.
TAP’s engine consists of a dimer, with two similar but nonidentical copies of a nucleotide binding domain, each of which functions like a cylinder. Each has a binding site for the cell’s fuel, ATP, and converts ATP into usable energy by hydrolyzing it. But, the asymmetrical sequence of the linear polypeptides foretold that one cylinder may be defective and hydrolyze ATP inefficiently. Gaudet was interested in why that sequence asymmetry exists, and her lab has been studying the matter through biochemistry and structure.
Their findings, she explains, will soon be ready for publication and could impact fields beyond the study of MHCs and immunology, including cystic fibrosis and cancer therapy. In CF, thick, salty mucous builds up in the lungs and harbors bacteria. It turns out that the intracellular portion of the protein that transports chloride through the cell membrane bears a homologous shape to TAP’s asymmetric dimer. Mutated forms of this protein affect the lung’s secretions. Several similarly structured transporter proteins excrete toxins in the liver. Some cancer cells produce more of these proteins to purge anticancer drugs, creating drug resistance.
"Having the structure of one of these proteins helps us think about how all the proteins from this family of transporters work and how to design better drugs against them," Gaudet says.
When setting up her own lab, Gaudet says, "I was looking for a family of proteins with lots of information to gain, not just small increments, where we could learn something fundamental about how the whole family works from looking at their structure."
She chose the large family of temperature-sensing proteins called TRPs as her inaugural project, which Strominger calls a "very original and very difficult problem." TRPs form ion channel proteins, several of which are expressed in the pain receptors (nociceptors) of peripheral nerves. "They evolved to let the brain know about the temperature of things," says Gaudet, "and they respond to temperature in ways never seen before in other proteins."
Understanding how these proteins work could have important medical implications, both for understanding different types of pain and for relieving it. But the same proteins are also involved in other functions, including taste and pheromone perception in mice. One family member, TRPV1, senses both types of heat, that from fire and that from the capsaicin in chili pepper. Another, TRPM8, senses cold temperature and is activated by minty menthol. TRPA1 responds to chemicals that generate pain and also to compounds in wasabi, mustard, garlic, and cinnamon.
"Once you study TRPs, you never think about food in the same way!" Gaudet remarks. "And they make great projects for undergraduate students, because they can rapidly relate to these proteins."
Recently, Gaudet made progress with TRPV2, which senses higher temperatures than TRPV1. Again, she first tackled the intracellular component of this protein, which appeared to contain three ankyrin repeat sequence motifs. Ankyrin repeats typically bind to other proteins, and the assumption was that these ankyrin repeats would join together the four subunits as a tetramer to create the actual ion channel. But when Gaudet succeeded in resolving the structure of this domain, she found six, not three, ankyrin repeats and learned that they do not form a tetramer after all.
"We published at the end of June, and right away I got emails from other researchers saying this study is really cool and might apply to a lot of other proteins," Gaudet says. She is pursuing a few clues in the hope of learning what the TRPV2 ankyrin repeats actually attach to. But it’s valuable to know that sequences alone cannot yet predict protein structure, she contends: "The people who work with sequences can use this knowledge to make better predictions. We can help each other out."
Gaudet next wants to discover what it is about heat that activates the TRP proteins. By studying proteins that sense different temperatures, she hopes to better understand general temperature sensing, both painful and non. She also plans to focus on the transmembrane portions of both TRP and TAP proteins – a daunting task. She’s also beginning a collaboration with Nancy Andrews at Harvard Medical School on proteins that transport divalent metals such as iron and manganese, which are used for energy metabolism and oxygen transportation.
Meanwhile, Gaudet is making a mark on undergraduate graduation, according to Strominger. She is now the co-head tutor for Biochemical Sciences with Richard Losick.
Gaudet says the diversity of MCB is a great asset to her. She is grateful for the support that the department gave her during her first foray into neuroscience with her work on pain receptors and temperature sensing. The neuroscience community at large has given her a vote of confidence, awarding her with the McKnight Scholar Award in the Neurosciences in 2004 for her project on TRP channels. She is also relieved about MCB’s vote of confidence in promoting her to Associate Professor: "Now I can get on with the research" – and on with the dancing.
Shortly after finishing this interview, Gaudet was off with her T Token Swing partner to the Camp Hollywood competition in Los Angeles. But in spite of her sizzling stage persona, she knows where her priorities lie. Last fall she achieved her biggest artistic accomplishment so far, qualifying for the World Lindy Hop Championships. "I couldn’t go, though," she says stoically, "because of a grant deadline."