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Expanding the genetic code: the world’s first truly unnatural organism

14.01.2003


Expanding the genetic code: the world’s first truly unnatural organism


From time immemorial, every living thing has shared the same basic set of building blocks - 20 amino acids from which all proteins are made.


That is, until now: A group of scientists say they have, for the first time, created an organism that can produce a 21st amino acid and incorporate it into proteins completely on its own. The research should help probe some of the central questions of evolutionary theory.

The findings are scheduled to appear in the Jan. 29 print edition of the Journal of the American Chemical Society, a peer-reviewed journal of the world’s largest scientific society. The article was initially published Jan. 4 on the journal’s Web site.



The project could eventually give concrete answers to questions that have generally been regarded as purely speculative: Is 20 the ideal number of basic building blocks? Would additional amino acids lead to organisms with enhanced function? Why has the genetic code not evolved further?

"Why did life settle on 20 amino acids?" asks Ryan Mehl, Ph.D., previously a researcher at the Scripps Research Institute in La Jolla, Calif., and now on the faculty of Franklin & Marshall College in Lancaster, Pa. "Would more amino acids give you a better organism -- one that could more effectively adapt if placed under selective pressure?"

To address this question, Mehl and a team of scientists led by Peter Schultz, Ph.D., professor of chemistry at Scripps, added a pathway to an E. coli bacterium that allows it to make a new amino acid -- p-aminophenylalanine (pAF) -- from simple carbon sources. Analytical techniques showed that pAF was incorporated into proteins with a fidelity rivaling that of the 20 natural amino acids.

"This allows you to have a totally autonomous organism that you can ’race’in one pot by evolving the new bacterium alongside its ancestors with 20 amino acids," says Christopher Anderson, a researcher at Scripps and another author of the paper. By racing the organisms -- exposing both to selective pressures at the same time and watching their development -- the researchers hope to see if the organism with the expanded genetic code has an evolutionary advantage over natural organisms.

A number of scientists have previously added unnatural amino acids to organisms, but most of these experiments involved eliminating the organism’s supply of the natural amino acid and substituting a close relative. "So, in the end, you still have a 20 amino acid bacterium, but it’s using an unnatural amino acid instead of the natural one," Anderson says.

"What our group really wanted to do is expand the genetic code, not just recode it. To do that, it takes a lot more effort. You have to come up with some way of specifically denoting how the protein is going to encode this 21st amino acid, because everything else in the genetic code already has a meaning associated with it."

To solve the problem, they used a process called amber suppression. This requires taking a stop codon -- a chunk of the genetic code that acts as a roadblock for protein synthesis -- and making it no longer mean "stop." Instead it now codes for the unnatural amino acid, so that the only way to suppress the codon is with the proper unnatural amino acid. "So you basically have a whole new pathway that you’ve created where the unnatural amino acid gets specifically [placed] onto a t-RNA," Anderson says.

The true novelty of the current paper is in biosynthesis -- the ability of the bacterium to make the new amino acid by itself, as opposed to being fed an unnatural amino acid from an outside source. "This bug is self-sufficient; it can make, load and incorporate the new amino acid in the emerging protein all on its own," Mehl says. "It’s a bona fide unnatural organism now. Essentially, this bacterium can be added to a minimal media (salts and a basic carbon source) and it’s able to do the rest."

E. coli is notorious for its ability to quickly reproduce, which could conjure images of mutant bacteria running wild. "We crippled the organism’s ability to biosynthesize leucine [one of the 20 essential amino acids] to avoid any risk that the organism could propagate outside a controlled lab setting," Anderson says. "Our unnatural organism will always live in the lab. We have no intention of putting it out in the wild or in commercial products where it could ’get out.’"

How this organism behaves in future experiments will determine, in part, where the research goes from here. "We are now focusing on more ’useful’ unnatural amino acids such as ketone- and PEG-containing amino acids," Anderson says. PEG stands for polyethylene glycol, a polymer that can be connected to proteins used in medicines to enhance their therapeutic value. "I don’t think it is at all unrealistic to imagine that in the not-too-distant future there will be a transgenic goat that can biosynthesize a PEG amino acid and incorporate it into therapeutic proteins secreted into the animal’s milk," Anderson says. "We are just beginning to look at the applications, but we have many projects in the works."

Contact: Beverly Hassell
e-mail: b_hassell@acs.org

Beverly Hassell | EurekAlert!

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