I chose aminoacyl-tRNA synthetases (aaRSs) to study today because these enzymes seemed like some of the more complex and mysterious molecules from this week’s lectures on protein synthesis. The more I researched, the more I understood how one could completely dedicate one’s life to studying only one enzyme. I read an in-depth paper by Rubio Gomez and Ibba (CSHL Press) and was surprised to discover that there are currently 23 known aaRSs. In addition to the 20 that Goodsell references that each code for a specific amino acid, there are two that code for lysine and two called pyrrolysyl-tRNA synthetase and phosphoseryl-tRNA synthetase found in some archaea and bacteria (Goodsell) (Rubio Gomez and Ibba).
Interestingly, aaRSs presents some interesting questions regarding their evolution, and, as others have mentioned, there are rare genetic disorders linked to aaRs mutations, as well as recent aaRs-targeted drug developments. Interestingly, these enzymes play a central role in the latest biosynthetic research as engineered amino acids are being written into novel polypeptide chains, altering the genetic code and resulting in the creation of biosensors, biomarkers, innovative functioning proteins, viral defenses, and more. Scientists have altered some aaRs:tRNA pairing to associate a new amino acid into translation (Rubio Gomez and Ibba) (Rovner et al.). As there is so much to discuss regarding aminoacyl-tRNA synthetases, I will focus on how they evolved and how they are altering evolution.
AaRSs are antiquated, having been inherited from the Last Universal Common Ancestor (LUCA) (Fournier et al.). And, they seem to have been just as complex in LUCA as they are in contemporary organisms (Rubio Gomez and Ibba). Since aaRSs are the readers of the genetic code, but also the genetic code is required to synthesize them, this presents a ‘chicken or the egg’ dilemma (Rubio Gomez and Ibba). As discussed in lecture 39, there are two types of aaRSs, each approaching the tRNA from a different side with their active site, with type I adding an amino acid to the last 2’ tRNA hydroxyl group and type II adding an amino acid to the 3’ hydroxyl on the final tRNA base (Barbaro). Both types also have a difference in their substrate binding methodology. Despite their difference, there is an accepted theory called the Rodin and Ohno hypothesis that both classes arose from the same gene simultaneously from opposing sides. The codons that code for residues for class I active sites are palindromes for class II sites. In other words, codons for class I are anticodons for class II. (Martinez-Rodriguez et al.). And so, two aaRSs arose due to bidirectional reading of mRNA and attached two different amino acids to tRNAs. This created the first protein comprised of more than one amino acid. Later genetic mutation and editing enabled the diversity of present-day aaRSs (Rubio Gomez and Ibba).
Noncanonical amino acids (ncAAs) are artificially synthesized amino acids that alter the genetic code and create genomically recoded organisms (GROs). GROs are created when scientists reassign a codon to a different and sometimes artificially synthesized amino acid (Lajoie et al.). Thus, new aaRSs are required that can recognize ncAAs and attach them to tRNAs that will transport them to ribosomes for protein chain synthesis. For example, Rovner et al. conducted research on an organism that lacked the TAG codon and reassigned TAG to code for a Methanocaldococcus jannaschii tRNA:aminoacyl-tRNA synthetase pairing. The protein was fabricated and stable.
I was curious to understand how aaRSs are altered to bind nCAAs. I came upon a study that described exactly its methodology and provided a helpful infographic. If you take a look at part c 2. (top right), you can see that a library of aaRS variants is generated after the introduction of the novel amino acid. Through mutation, an aaRS will mutate to associate the codon with the ncAA. This will be selected and amplified using PCR technology (Vargas-Rodriguez et al.). I was surprised that ncAAs are created by selective breeding!
Image: www.ncbi.nlm.nih.gov/pmc/articles/PMC6214156.
I have only scratched the surface of my understanding of aaRSs. From ancient origins to the future of genetic engineering, it seems there is ad infinitum to discover in the study of aminoacyl-tRNA synthetases.
REFERENCES
Rubio Gomez, Miguel Angel, and Michael Ibba. “Aminoacyl-tRNA synthetases.” RNA (New York, N.Y.) vol. 26,8 (2020): 910-936. doi:10.1261/rna.071720.119
Fournier, Gregory P., et al. "Molecular Evolution of Aminoacyl tRNA Synthetase Proteins in the Early History of Life." Origins of Life and Evolution of Biospheres, vol. 41, no. 6, 2011, pp. 621-632.
Barbaro B, Biochemistry at U of California San Diego. Course Number: Chapter 39 - the Genetic Code” [accessed 2023 Dec 15]
Martinez-Rodriguez, Luis, et al. “Functional Class I and II Amino Acid-activating Enzymes Can Be Coded by Opposite Strands of the Same Gene.” PubMed Central (PMC), 18 June 2015, www.ncbi.nlm.nih.gov/pmc/articles/PMC4528134 Links to an external site..
Rovner, Alexis J., et al. “Recoded Organisms Engineered to Depend on Synthetic Amino Acids.” PubMed Central (PMC), 21 Jan. 2015, www.ncbi.nlm.nih.gov/pmc/articles/PMC4590768 Links to an external site..
Goodsell, David. “PDB101: Molecule of the Month: Aminoacyl-tRNA Synthetases.” RCSB: PDB-101, pdb101.rcsb.org/motm/16. Accessed 16 Dec. 2022.
“Genetically Modified Organisms (GMOs) | Learn Science at Scitable.” Genetically Modified Organisms (GMOs) | Learn Science at Scitable, www.nature.com/scitable/topicpage/genetically-modified-organisms-gmos-transgenic-crops-and-732. Accessed 16 Dec. 2022.
Lajoie, Marc J., et al. “Genomically Recoded Organisms Expand Biological Functions.” PubMed Central (PMC), www.ncbi.nlm.nih.gov/pmc/articles/PMC4924538. Accessed 16 Dec. 2022.
Vargas-Rodriguez, Oscar, et al. “Upgrading aminoacyl-tRNA Synthetases for Genetic Code Expansion.” PubMed Central (PMC), 27 July 2018, www.ncbi.nlm.nih.gov/pmc/articles/PMC6214156.