The genetic code is the recipe for life, and provides the instructions for how to make proteins, generally using just 20 amino acids. But certain groups of microbes have an expanded genetic code, in which one or two additional amino acids are inserted into the protein — a finding that has been leveraged for bioengineering. In a new paper out today in Science, Veronika Kivenson, working with IGI Investigator Jill Banfield, UC Berkeley’s Alanna Schepartz, and collaborators at CGEM, Institut Pasteur, and elsewhere identify a new genetic code present in multiple kinds of microbes called archaea. The findings may help scientists reduce greenhouse gas emissions, and could also be exploited by chemists to create novel proteins and useful polymer-like materials.
Genes are made of codons — that is, sequences of three DNA or RNA letters that specify an amino acid or stop signal during protein synthesis. Expanded genetic codes typically repurpose one of the canonical stop codons to instead trigger the addition of an unusual amino acid. There have previously been rare examples in archaea in which a stop codon is instead used to add the amino acid pyrrolysine. But until now, this event was only known to occur naturally in a limited number of specific proteins in specific branches of the tree of life. In a new paper, the researchers identify several groups of archaea that read the stop codon in question as a pyrrolysine signal — every time.
“An expanded genetic code is one in which the DNA is read in a different way than expected. Until now, only one genetic code was ever reported in archaea but this paper reports a new genetic code. The particular stop codon is now universally read as pyrrolysine,” says Banfield. “Only if we understand the code can we understand these organisms and what they can do.”
This new expanded code has arisen many times at different places in the archaeal genetic tree, challenging the conventional view that genetic code changes are rare events.
Understanding & Intervening in Methane Cycles
Many of the archaeal proteins that use pyrrolysine are involved in the cycling of methane, a greenhouse gas 27x more potent than carbon dioxide and a major contributor to global warming. In fact, all of the archaea with the expanded genetic code are known to produce methane. Evolution of this alternative genetic code was driven by a metabolic need for pyrrolysine-containing enzymes that enable consumption of compounds called methylamines, which are common in the environment. The methyl part of these compounds is converted to methane.
Prior to the team’s findings, the genetic code of these microbes was misinterpreted: scientists couldn’t see hundreds of proteins containing pyrrolysine. Correctly identifying this code now enables improved understanding of the roles of these organisms in methane cycles and greenhouse gas emissions.
Creating Novel Proteins for Medicine & More
Genetic code expansion is used widely for fundamental science research and bioengineering. It was first harnessed for creating novel proteins almost 25 years ago, by then UC Berkeley professor Peter Schultz. Schultz was able to reprogram a stop codon to introduce amino acids that don’t naturally occur in cells to make bespoke proteins.
Microbes with expanded codes can be grown as purified cultures, or components of their genomes can be put directly into E. coli bacteria, yeast, or human cells, working as protein factories contained within the cells.
Based on Schultz’s research, hundreds of different new-to-nature amino acids have now been engineered into proteins using genetic code expansion. These proteins have been used in research and in industry to make antibody medications and immunotherapies.
What the New Code Can Offer
The Banfield lab team wondered if their findings could be used for bioengineering, so they turned to UC Berkeley chemist and synthetic biologist Alanna Schepartz.
Schepartz and her team tested out pyrrolysine systems isolated from eight different groups of archaea by putting them in the bacteria E. coli. These E. coli were programmed to make a fluorescent protein with the stop codon in question in the middle of the protein sequence. Without the pyrrolysine system, the protein would be only half-made and unable to fluoresce. But the pyrrolysine systems all worked as predicted, interpreting the codon as an amino acid directive, not a stop sign, and making the bacteria glow.
“The big challenge is that genetic code expansion is not always predictable,” says Schepartz. “The success of the current technology varies depending on the RNA sequence context, the microbe strain, and/or the structure of the new-to-nature amino acid. It’s very hard to predict success, so a lot of experimentation is needed to optimize a system to ensure high yields and purity. The organisms identified in this work appear to have figured out a natural solution to this problem. It’s amazing, and yet another fantastic example of how biology hides secrets that drive biotechnology innovation.”