When T. rex is part of a day’s work John Asara’s journey into analytical chemistry and proteomics leads him to dinosaurs.
got his first chemistry set when he was onduct a Google search for John eight years old. Children’s chemistry Asara, and you’ll probably think sets, Asara notes, were less regulated in his he’s a dinosaur hunter. Over the past two years, the researcher at Harvard Medical School and Beth Israel Deaconess Medical Center (BIDMC) has been inextricably linked to a 68-million-year-old Tyrannosaurus rex in media outlets and peer-reviewed journals. Asara is bemused by the skewed view the world has of his work: he considers himself a regular analytical chemist. At the MS facility Asara directs at BIDMC, his day is largely focused on collaborative projects with other scientists and his own research in cancer biology. The facility specializes in A collaboration with Mary Schweitzer on a 68-milliontargeted and quantitative proteomics year-old Tyrannosaurus rex catapulted John Asara’s with emphasis on using LC/MS/MS name into headlines. to find protein⫺protein interaction networks and posttranslational modifica- day than they are now, so he spent some tions involved in cellular signal-transof his youth playing with alcohols, Bunsen duction pathways. He spends at most burners, and lead-containing salts. 20% of his time on the dinosaur analysis, Chemistry continued to dominate he says. Asara’s interests when he enrolled at Because he welcomes all kinds of colBrandeis University for his undergradulaborations, Asara is “not tethered to a ate degree and later at Michigan State specific area,” explains Jeffrey Engelman, University (MSU) for his Ph.D. At who is at Harvard Medical School and Brandeis, he became interested in anaMassachusetts General Hospital and has lytical chemistry because “I recognized collaborated with Asara for several years the influence of analytical chemistry in on cancer research. “He has a natural the biological sciences,” he says. “I realcuriosity about biology and a very ized that was the way things were movstrong desire to see how to best utilize ing and what I had to get into.” MS to advance different biological Asara hadn’t chosen an area of spefields.”. cialization when he arrived at MSU in It was this natural curiosity of Asara’s 1995. But he came across the MS labothat got him involved four years ago in ratory of John Allison, who is now at the the dinosaur project with Mary SchCollege of New Jersey. “I recognized weitzer at North Carolina State that MS was becoming a bigger influUniversityOand thrust him into the ence on the biological sciences,” says glaring spotlight of media attention. Asara. Besides, mass spectrometers
Chemistry sets to mass spectrometers Asara’s foray into the field was typical of many who ended up being scientists. He 8660
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“were big instruments and looked fun to use!” Asara worked on fast-atom bombardment and MALDI MS, work that he credits for teaching him the fundamentals of MS.
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One of his projects was to develop matrix additives to help find molecules that usually eluded detection. His particular interest was phosphorylated peptides. Allison tells his favorite anecdote about his former student: Asara came across a commercially available kit for detecting phosphorylated peptides. “He bought the kit, developed his own approach, and showed that he could do much better than what was commercially available,” Allison recalls. Then, laughs Allison, Asara sent a note to the vendor of the kit explaining how to improve its product! When Asara graduated with his Ph.D. in 2000, he joined William Lane’s laboratory at Harvard University. Lane runs a successful MS proteomics core facility, and that’s where Asara cut his teeth on proteomics. A brief stint at a biotech firm followed his postdoctoral positionOuntil he saw a job posting for someone to run an MS proteomics facility at BIDMC.
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BIDMC In 2004, Lewis Cantley at BIDMC was looking for someone to build an MS core facility from scratch. “All we had was money to buy an instrument!” chuckles Cantley. During the interview, Asara impressed Cantley. “We didn’t want a person to just run a state-of-the-art facility and do a 9-to-5 job,” says Cantley. “We wanted someone who’d keep us at the frontier of the field.” Asara was hired and given a corner in Cantley’s laboratory. Soon, Asara got funding through grants, garnered local resources, bought instruments, and had the MS facility up and running. Today, the core facility is independent of Cantley’s lab and boasts several MS instruments, including a fast-scanning 2D linear ion trap, an LTQ Orbitrap XL ETD,
10.1021/AC9020215 2009 AMERICAN CHEMICAL SOCIETY
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and a 5500 QTRAP mass spectrometer designated for metabolomics research. Asara and his team provide services in protein identification, posttranslational modification mapping, and relative peptide/protein quantification by stableisotope labeling and label-free methods based on microcapillary LC/MS/MS techniques. Cantley says the facility’s success reflects Asara’s work ethic. “At most MS cores, you hand off the sample at the door, and three weeks later, you get an email with the result. If you want to talk to somebody, it’s someone at a technician level,” he says. “John isn’t like that. People come to him because he has the reputation for having the instrumentation to tackle difficult questions and being willing to engage with people and spend hours on the phone discussing what they are trying to get out of the experiment and what would be the best approach.”
Stepping onto dino-mite In 2005, as the MS facility grew, Cantley happened upon a newspaper story. Schweitzer had just published molecular analyses of soft tissues from a 68-million-year-old T. rex (Science 2005, DOI 10.1126/science.1108397). Schweitzer wasn’t a stranger to Asara. She had previously collaborated with Lane, Asara’s postdoctoral adviser, on fossil analysis. And Asara wasn’t a stranger to the study of fossils: during his postdoctoral training, he had worked on a project to analyze protein from a 100,000⫺300,000year-old mammoth (J. Mol. Evol. 2002, DOI 10.1007/s00239-002-2365-6). At that time, it was the oldest protein sequenced. “I was impressed by the sensitivity of the instrumentation we had acquired, which was much more powerful than instruments that existed five years earlier,” explains Cantley. “When I read in
the New York Times about identification of T. rex bones that appeared to have structures that looked like blood vesselsOraising the possibility it was well-preservedOI jokingly sent off an email to John, saying, ‘I think you can beat your own record of a 100,000year-old protein!’” Asara approached Schweitzer for a collaboration. Schweitzer agreed, but even before extracts passed from her laboratory to his, she had some advice for him. Schweitzer “definitely warned me about the attention the research would get, and the criticism, even before we had data!” says Asara. When the two teams published their analyses in 2007 in Science stating that they had discovered collagen-like peptides that closely matched bird collagen, criticism of the work began to spread in the journals and media. The criticism became particularly explosive when Pavel Pevzner at the University of California San Diego published a stinging critique, saying the analyses were statistically insignificant. (See the AC news story “Uproar over dinosaur data” and Wired magazine’s story.) Despite Schweitzer’s warnings, Asara admits, “I wasn’t prepared for the impact [the paper] would have in the media and the criticism by other people in the field.” He had been raked over the coals in publicOand he felt that way. His shock, however, wasn’t apparent to people like Engelman, who watched Asara during the T. rex controversy. Engelman thought Asara seemed upbeat, even then. “He’s more on the optimistic side of the world than the pessimistic,” says Engelman. To defend the T. rex work, Asara and Schweitzer followed up with an analysis of an even older dinosaur fossil; this helped to calm the storm to some extent (see AC story, “New analysis seen as validating dinosaur proteomics study”).
Asara says he learned a lot from the experience, “because every step I took became a public issue.” Even Asara’s attempt to meet one of Pezvner’s demandsOthat Asara release the entire T. rex data set of mass spectra into a public databaseObecame a controversy. Asara initially released the spectra with a stipulation limiting how the data could be used, an unprecedented move that unleashed more criticism. (See the Journal of Proteome Research news story, “A controversial data set stirs up even more controversy”.) That experience has given Asara a new respect for the public release of raw data. Now he thinks a data set as highprofile as the T. rex one benefits from the scrutiny of other scientists. In this case, in fact, other scientists have revisited this data set and published findings that support the original paper. (See the Journal of Proteome Research story, “Independent analysis of controversial T. rex data confirms findings”.)
The dinosaurs stay These days, Asara is busy cranking through his cancer proteomics projects. At the Human Proteome Organisation VIII World Congress in Toronto this year, for instance, he and his colleagues presented work in which they teased out the interactome of parts of the insulin signaling pathway in fruit flies by immunoprecipitation and MS. They carried out similar studies on human cell lines of myeloma non-small-cell lung cancer and, by comparing the fly and human data, discovered several novel members of the pathway. But despite the trials and tribulations of the T. rex studyOand the fact that the bulk of his work is focused on cancer biologyOAsara seems hooked on dinosaurs. He says, “I consider part of my lab to be dedicated to ‘fossilomics’.” —Rajendrani Mukhopadhyay
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