Scientists add new letters to bacteria’s genetic ‘alphabet’

For possibly billions of years, the DNA blueprints for life on Earth have been written with just four genetic “letters” -- A, T, G and C. On Wednesday, scientists announced that that they added two more.

In a paper published in the journal Nature, bio-engineers at Scripps Research Institute in La Jolla said they had successfully inserted two synthetic molecules into the genome of an Escherichia coli bacterium, which survived and passed on the new genetic material.

In addition to the naturally occurring nucleotides adenine, thymine, guanine and cytosine, which form the rungs of DNA’s double-helix structure, the bacterium carried two more base-pair partners, which study authors have dubbed d5SICS and dNaM.

For more than a decade, scientists have been experimenting with so-called unnatural base pairs, or UBPs, saying they may hold the key to new antibiotics, future cancer drugs, improved vaccines, nanomaterials and other innovations.

Until now, however, those experiments have all been conducted in test tubes.

“These unnatural base pairs have worked beautifully in vitro, but the big challenge has been to get them working in the much more complex environment of a living cell,” lead study author Denis Malyshev, a molecular and chemical biologist at Scripps, said in a prepared statement.

The new genetic material did not appear to be toxic to the bacteria, and it only remains in the organism’s genome under specific lab conditions. In a natural environment, the molecules -- nucleoside triphosphates -- degrade and disappear in a day or two. Once they disappear, the bacterium reverts back to its natural base pair arrangement.

Still, experts said insertion of the synthetic materials into E. coli’s genome was a milestone.

“This definitely is a significant achievement,” said Ross Thyer, a synthetic biologist at the University of Texas at Austin, who was not involved in the research. “What I’m most excited about is how this will help us answer some bigger evolutionary questions: Why has life settled on a specific set of bases.”

Malyshev and colleagues went about creating the semi-synthetic bacterium by genetically engineering a stretch of ring-like DNA known as a plasmid.

The engineered plasmid contained E. coli’s usual complement of coordinated A,T, G and C nucleotides, as well as two man-made molecules, which join to form a new rung on the DNA ladder.

But the task of getting bacteria to maintain those molecules in their DNA was far more difficult.

Like all genetic material, the new molecules degrade over time. Although cells routinely repair their naturally occurring nucleotides with materials on hand, the E. coli have no means of producing the foreign synthetic materials.

If this man-made genetic material was to survive within the bacteria and be passed on during reproduction, the study’s authors reasoned that they would have to surround the cells with a solution containing the new material. They would also have to create a doorway through which the synthetic molecules could enter the cell.

To create this portal, study authors engineered an E. coli strain that expressed an algal nucleotide triphosphate transporter (NTT) protein, which would recognize the necessary molecules in a surrounding medium and escort them into the cell.

“This was the big breakthrough for us,” Malyshev said.

Once those conditions were met, the semi-synthetic bacteria survived and appeared to reproduce without “a notable growth burden,” authors wrote. Also, the cell did not attack and remove the foreign material.

“Thus, the resulting bacterium is the first organism to propagate stably an expanded genetic alphabet,” authors wrote.

The next step for synthetic biologists will be to give experimental cells a reason to want to keep the synthetic genetic coding.

“Right now, if they’re gradually lost, the cells don’t care,” Thyer said. “So one of the next big challenges is to engineer the cells to become dependent on these unnatural bases. We need to give them some function that provides a significant benefit to the cells.”