Genetic coding revamp

Scientists have developed a new genetic language using a ribosome that can read instructions that are 4 base pairs long, enabling the construction of designer proteins containing a variety of unnatural elements, according to a study published online today (February 14) in Nature.

Image: Wikimedia commons,
S. Jähnichen

"It is brilliant," said organic chemist linkurl:John Sutherland;http://www.chemistry.manchester.ac.uk/aboutus/staff/showprofile.php?id=390 of the University of Manchester in the UK, who was not involved in the work. "[It's] a whole new genetic code [that can be used to create] all sorts of different polymers with all sorts of applications. It's a truly important piece of science." For years, scientists have been working to incorporate what they term unnatural amino acids -- structurally modified amino acids with different physicochemical and biological properties -- into proteins. The aim is to better understand normal cellular processes, such as the functional effects of posttranslational modifications, as well as to develop artificial biological systems with novel properties. Most of the success in this endeavor has stemmed from the use of a single stop codon, known as the amber codon, that researchers have been able to manipulate to integrate modified amino acids during protein synthesis. By artificially evolving transfer RNA (tRNA) molecules that respond to the amber codon's UAG sequence, and linking modified amino acids to those tRNA molecules, scientists have been able to generate proteins with a variety of unnatural units. But with just one spare codon to work with, scientists have largely been restricted to incorporating only one such unit per protein. To overcome this limitation, synthetic biologist linkurl:Jason Chin;http://www2.mrc-lmb.cam.ac.uk/group-leaders/a-to-g/j-chin of the Medical Research Council Laboratory of Molecular Biology in Cambridge, UK, and his colleagues decided to devise a system that could read codons that are 4 base pairs long. Such a system could "open the door to what will be [the] truly revolutionary possibility [of creating] genetically coded polymers comprised of up to 256 [unique] building blocks." The problem was that normal ribosomes -- which translate 3-base codons into any of the 22 naturally occurring amino acids -- don't read such quadruplet codons, at least not usually. Furthermore, the team couldn't simply manipulate the existing ribosomes, as they are responsible for producing all of the cell's proteins that are required for its survival; altering this system could quickly cause a cellular collapse. Their solution: make a whole new ribosome. A few years ago, the researchers linkurl:created the new ribosome,;http://www.nature.com/nchembio/journal/v1/n3/full/nchembio719.html known as an orthogonal ribosome, by altering the region that recognizes the ribosome-binding sequence of the messenger RNA (mRNA). They then created special mRNAs with complementary binding regions to this new sequence, which the orthogonal ribosome selectively bound to and read, leaving natural mRNAs to be recognized only by the natural ribosomes. "Now you've got two ribosomes -- one reading a new message and [the] normal ribosome" reading the old messages, Chin explained. "That's the basis of how you would write a parallel genetic code in the cell." With the two ribosomes systems working independently, the researchers could then manipulate the orthogonal ribosome without disrupting normal cellular function. In the present study, they did just that, inducing mutations in the ribosome where the tRNA and mRNA molecules interact in hopes of creating a ribosome that could read quadruplet codons with comparable efficiency and accuracy to that of natural protein synthesis. To test their mutated ribosomes, the team put them in bacterial cultures growing on a medium containing antibiotics, and provided the cells with an antibiotic resistance gene that included a 4-base codon. Ribosomes that could read the quadruplet codon successfully produced the antibiotic resistance protein, and survived even in the presence of high concentrations of the antibiotic. Those that couldn't read the quadruplet, couldn't create the protein to protect themselves from the antibiotic and died as a result. "In the end we get cells that are surviving this selection pressure," Chin said, and in those cells are ribosomes that can successfully read quadruplet codons. The new ribosome can still read triplet codons as well, Chin added, but it preferentially reads the 4-base sequences. In this way, the ribosome can still incorporate natural amino acids as well as modified units attached to the amber codon. But now with the ability to read the quadruplet codons, scientists can easily create proteins with more than one unnatural unit. Using the amber codon and a quadruplet codon to code for two different unnatural amino acids, the researchers generated a protein, synthesized by the new ribosome, that contained both unnatural units. "I think it's fantastic," Sutherland said. "It opens up an era of synthetic biology where we are truly making synthetic polymers," as opposed to just "tweaking and tinkering" with an existing protein. "I believe this to be a milestone contribution to the protein chemistry field," linkurl:Tom Muir;http://www.rockefeller.edu/research/faculty/abstract.php?id=121 of The Rockefeller University, who did not participate in the research, agreed in an email to The Scientist. "This could open the floodgates to all manner of exciting applications."
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[10th October 2005]