Journal Article10.1021/acs.chemrev.3c00878
Cracking the Code: Reprogramming the Genetic Script in Prokaryotes and Eukaryotes to Harness the Power of Noncanonical Amino Acids
Cosimo Jann,Sabrina Giofrè,Rajanya Bhattacharjee,Edward A. Lemke +3 more
4
TL;DR: Researchers review genetic code expansion methodologies in prokaryotes (E. coli) and eukaryotes (mammalian cells), highlighting challenges and promising developments in harnessing noncanonical amino acids for biotechnology, materials science, and medicine applications.
read more
Abstract: Over 500 natural and synthetic amino acids have been genetically encoded in the last two decades. Incorporating these noncanonical amino acids into proteins enables many powerful applications, ranging from basic research to biotechnology, materials science, and medicine. However, major challenges remain to unleash the full potential of genetic code expansion across disciplines. Here, we provide an overview of diverse genetic code expansion methodologies and systems and their final applications in prokaryotes and eukaryotes, represented by Escherichia coli and mammalian cells as the main workhorse model systems. We highlight the power of how new technologies can be first established in simple and then transferred to more complex systems. For example, whole-genome engineering provides an excellent platform in bacteria for enabling transcript-specific genetic code expansion without off-targets in the transcriptome. In contrast, the complexity of a eukaryotic cell poses challenges that require entirely new approaches, such as striving toward establishing novel base pairs or generating orthogonally translating organelles within living cells. We connect the milestones in expanding the genetic code of living cells for encoding novel chemical functionalities to the most recent scientific discoveries, from optimizing the physicochemical properties of noncanonical amino acids to the technological advancements for their in vivo incorporation. This journey offers a glimpse into the promising developments in the years to come.
read more
Chat with Paper
AI Agents for this Paper
Find similar papers on Google Scholar, PubMed and Arxiv
Write a critical review of this paper
Analyze citations of this paper to find unaddressed research gaps
Citations
Non-canonical amino acids for site-directed spin labeling of membrane proteins
Kaitlyn Ledwitch,Georg Künze,Elleansar Okwei,Davide Sala,Jens Meiler +4 more
TL;DR: Researchers utilize non-canonical amino acids to develop site-directed spin labeling methods for membrane proteins, enabling atomic-level structural and dynamical information through nuclear and electron paramagnetic resonance applications in complex lipid environments.
Noncanonical Amino Acid Incorporation in Animals and Animal Cells
Joo‐Chan Kim,YouJin Kim,Suho Cho,Hee‐Sung Park +3 more
TL;DR: Noncanonical amino acid incorporation technology enables precise control over biological processes in animals and animal cells, offering novel insights into human physiology, genetics, and diseases, with applications in model organisms and potential therapeutic interventions.
Hijacking a bacterial ABC transporter for genetic code expansion
Tarun Iype,Maximilian Fottner,Paul Böhm,Carlos Piedrafita,Yannis Möller,Michael Groll,Kathrin Lang,Tarun Iype,Maximilian Fottner,Paul Böhm,Carlos Piedrafita,Yannis Möller,Michael Groll,Kathrin Lang +13 more
Abstract: Abstract The site-specific encoding of non-canonical amino acids (ncAAs) provides a powerful tool for expanding the functional repertoire of proteins 1–4 . Its widespread use for basic research and biotechnological applications is, however, hampered by the low efficiencies of current ncAA incorporation strategies. Here we reveal poor cellular ncAA uptake as a main obstacle to efficient genetic code expansion and overcome this bottleneck by hijacking a bacterial ATP-binding cassette (ABC) transporter 5 to actively import easily synthesizable isopeptide-linked tripeptides that are processed into ncAAs within the cell. Using this approach, we enable efficient encoding of a variety of previously inaccessible ncAAs, decorating proteins with bioorthogonal 6 and crosslinker 7 moieties, post-translational modifications 8,9 and functionalities for chemoenzymatic conjugation. We then devise a high-throughput directed evolution platform to engineer tailored transporter systems for the import of ncAAs that were historically refractory to efficient uptake. Customized Escherichia coli strains expressing these evolved transporters facilitate single and multi-site ncAA incorporation with wild-type efficiencies. Additionally, we adapt the tripeptide scaffolds for the co-transport of two different ncAAs, enabling their efficient dual incorporation. Collectively, our study demonstrates that engineering of uptake systems is a powerful strategy for programmable import of chemically diverse building blocks.
Investigation of protein post-translational modifications with site-specifically incorporated non-canonical amino acids
Jiayu Gu,Lihui Lao,Yulin Chen,Shixian Lin +3 more
References
A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity.
Martin Jinek,Krzysztof Chylinski,Krzysztof Chylinski,Ines Fonfara,Michael H. Hauer,Jennifer A. Doudna,Emmanuelle Charpentier +6 more
TL;DR: This study reveals a family of endonucleases that use dual-RNAs for site-specific DNA cleavage and highlights the potential to exploit the system for RNA-programmable genome editing.
16.2K
Multiplex Genome Engineering Using CRISPR/Cas Systems
Le Cong,Le Cong,F. Ann Ran,F. Ann Ran,David M. Cox,David M. Cox,Shuailiang Lin,Shuailiang Lin,Robert P. J. Barretto,Naomi Habib,Patrick D. Hsu,Patrick D. Hsu,Xuebing Wu,Wenyan Jiang,Luciano A. Marraffini,Feng Zhang +15 more
TL;DR: The type II prokaryotic CRISPR (clustered regularly interspaced short palindromic repeats)/Cas adaptive immune system has been shown to facilitate RNA-guided site-specific DNA cleavage as discussed by the authors.
Click Chemistry: Diverse Chemical Function from a Few Good Reactions.
TL;DR: In this paper, a set of powerful, highly reliable, and selective reactions for the rapid synthesis of useful new compounds and combinatorial libraries through heteroatom links (C-X-C), an approach called click chemistry is defined, enabled, and constrained by a handful of nearly perfect "springloaded" reactions.
9.7K
Peptidotriazoles on solid phase: [1,2,3]-triazoles by regiospecific copper(i)-catalyzed 1,3-dipolar cycloadditions of terminal alkynes to azides.
TL;DR: A novel regiospecific copper(I)-catalyzed 1,3-dipolar cycloaddition of terminal alkynes to azides on solid-phase is reported, and the X-ray structure of 2-azido-2-methylpropanoic acid has been solved, to yield structural information on the 1, 3-dipoles entering the reaction.
8.3K