Tinkering with Tumor-tracking Tadpoles

By Edyta Zielinska

At the age of 14, Ian Burbulis blew off four fingers on his left hand during a pyrotechnics experiment gone wrong. But that didn't stop him. Burbulis was the kind of kid who, much to his mother's dismay, took apart all of his family's appliances just to peek inside. One time, he partly dismantled the timing dial of his mother's washing machine, so that he could watch how its parts moved, ticked, and triggered each consecutive cycle.

After working a few years at a construction company following high school, Burbulis landed scholarships to attend Virginia Tech. He started studying architecture but switched to biology, and then stayed to work on a PhD in molecular and cell biology. He knew how to take things...

"I decided to take the route of science and be the mechanic that I am," says Burbulis, who is soft-spoken with a slight Southern accent, and now has fingers of varying lengths on his left hand. "A lot of the things I want to understand demand a quantitative measure. I really want to understand how the engine works, to identify the parts that are broken so that I can reach inside the mechanism and replace it."

At the Molecular Sciences Institute, Burbulis and Rob Carlson began work on a tool that would let scientists peer into the cell and count, with precision, the number of molecules it contains. While detecting low levels of DNA has become simple with PCR technology, there was still no good way to detect minute quantities of protein. The best method to date was the enzyme-linked immunoabsorbent assay (ELISA), which gave only semiquantitative results. Using his mechanic's ingenuity and his biochemical knowledge, Burbulis decided that if he could link DNA to a particular protein, PCR could indirectly measure the amount of protein by amplifying the DNA to which it's attached. Thus, Burbulis' "Tadpoles" - with their protein heads and DNA tails - were born.

The trick was finding the right glue to stick the head and tail together. Using existing methods, Burbulis could produce only tiny and expensive quantities of the chimeric substance. Then he had a breakthrough. "I'm riding my bicycle to work one day, and I remembered the work of Fran Perler at New England Biolabs," says Burbulis. Perler discovered inteins, the self-splicing protein. Inteins were perfect for producing Tadpoles because, when bound to a protein, they excise themselves but leave behind a "hook," which DNA could bind to covalently. Burbulis engineered Escherichia coli to express his protein of interest along with an intein, and then mixed the protein with the DNA tails, which immediately bound the thioester hook.

This simplified reaction, which takes only a few hours of incubation followed by a few more hours for purification, allowed him to make the material en masse. "I'm talking about milligram or even gram quantities," said Burbulis.

With a sensitivity of 100,000 higher than the ELISA, and a relatively simple production scheme, Tadpoles (for which Burbulis has submitted a patent) can detect the exact numbers of molecules within the alpha-pheromone signaling pathway, and miniscule quantities of almost any other molecule. With clear applications for diagnostics and clinical trials, Burbulis was tapped by the National Cancer Institute's Innovative Molecular Analysis Technologies program, to develop Tadpoles for detecting low-level molecules in the blood that would be able to show cancer risk - work he's developing now.

Burbulis hasn't stopped being a mechanic and a lover of high octane. Sometimes he and another MSI scientist, Richard Yu, ride their motorcycles around Berkeley to take a break from their research. And, Burbulis is often asked to fix things in the lab. "I can't tell you how many times I've fixed that DNA sequencer," he says.

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