22nd amino acid synthesized and added to genetic code of e. coli bacteria

09/13/04

COLUMBUS, Ohio Two years ago, Ohio State University researchers surprised the scientific community by announcing their discovery of a 22nd genetically encoded amino acid.

Now they have capped that discovery with news that they have successfully synthesized the amino acid itself L-pyrrolysine and shown that bacteria can incorporate it into new proteins the biological components which do most of the work in cells.

The importance of their work is the explanation of exactly how the 22nd amino acid is incorporated into proteins inside living cells. The genetic instructions to put pyrrolysine into proteins follows a traditional path that many scientists had not predicted.

For decades following the discovery of the structure of DNA, the dogma was that the genes in the DNA were decoded to produce proteins built from only 20 "canonical" amino acids.

But in 1986, researchers discovered that a 21st amino acid selenocysteine was incorporated into certain proteins. What separated selenocysteine from the other previously identified amino acids was the fact that it was inserted into protein by a very different path.

Each of the canonical amino acids uses a specialized translator protein to decode genetic information as that amino acid. But selenocysteine lacked its own translator protein and is put into the protein through a more circuitous route.

That left open the question of whether future amino acids would follow the traditional path of the first 20 amino acids or the unusual route taken by 21st.

In the end, tradition won out.

The results are described in two papers published in scientific journals this month. The first appeared in the British journal Nature and the second in the journal Chemistry and Biology.

"In recent years, researchers learned to artificially modify a set of translator enzymes so that new amino acids can be genetically programmed in cells to produce novel proteins for biotechnology," explained Joe Krzycki, a professor of microbiology.

"Now it looks like nature has been doing the same kinds of experiments all along and has produced a never-before-seen translator protein for pyrrolysine."

Following their joint discovery of L-pyrrolysine, Krzycki's colleague Michael Chan, an associate professor of biochemistry and chemistry at Ohio State, began the laborious process of synthesizing the actual chemical compound. After nearly a year's worth of work and a 10-step process, Chan's research group provided Kyzycki's group with a small amount of this chemically synthesized pyrrolysine.

"The sample that Michael's team produced was exactly what the chemical structure that we'd predicted said it should be," Krzycki said.

The final step was to see if the translator enzyme for L-pyrrolysine would function normally within a living cell. To test that, they inserted the enzyme into Escherichia coli bacteria. Researchers use E. coli as an easy test for some basic biological functions.

Once inside the microbe, the enzyme was able to change the genetic coding in the organism so that it now included L-pyrrolysine as well as the other 21 amino acids.

"What this set of experiments has shown us is that it is probably a little easier than we might have thought it would be to change the genetic code in an organism like E. coli," Krzycki said. "It gives us some new insight into how the genetic code evolved, that's the take-home message."

Chan called the project itself "a great example of how researchers in chemistry and biology can come together to solve some important fundamental scientific questions."

Future research may suggest new ways to make artificial proteins with unusual chemical properties for use in medicine or industry, Krzycki said.

The research was supported by the National Science Foundation, the National Institutes of Health, the Department of Energy and the Alfred P. Sloan Foundation.

Along with Chan and Krzycki, Bing Hao, Gang Zhao, Patrick Kang, Jitesh Soares, Tsuneo Ferguson, Judith Gallucci, Sherry Blight, Ross Larue, Anirban Mahapatra, David Longstaff, Edward Chang, and Kari Green-Church all worked on the project.

Source: Eurekalert & others

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