“This work shows that enzymes are far more adaptable than we once thought,” says study leader John Chaput, UC Irvine professor of pharmaceutical sciences. “By harnessing evolution, we can create new molecular tools that open the door to advances in RNA biology, synthetic biology and biomedical innovation.”
Photo Credit: Steve Zylius / UC Irvine
Scientific Frontline: "At a Glance" Summary
- Main Discovery: Researchers engineered a novel DNA polymerase, designated C28, that efficiently synthesizes RNA with high fidelity and speed, a capability that natural DNA polymerases are biologically designed to reject.
- Methodology: The team utilized directed evolution within a high-throughput, single-cell screening platform to recombine related polymerase genes, evaluating millions of variants to identify unexpected structural solutions without manually redesigning the active site.
- Key Data: The C28 enzyme contains dozens of specific mutations selected from a pool of millions of variants, enabling it to operate at near-natural speeds while accommodating chemically modified RNA building blocks.
- Significance: This breakthrough overcomes fundamental biological barriers to RNA synthesis, creating a versatile tool that can also perform reverse transcription and generate hybrid DNA-RNA molecules using standard PCR techniques.
- Future Application: The enzyme provides critical functionality for developing next-generation mRNA vaccines and RNA-based therapeutics that require customized or chemically modified RNA sequences.
- Branch of Science: Biochemistry, Pharmaceutical Sciences, and Synthetic Biology.
- Additional Detail: Led by Professor John Chaput and published in Nature Chemical Biology, this research demonstrates that directed evolution can unlock molecular functions nonexistent in nature, such as the ability of a DNA polymerase to transcribe RNA.
From vaccines and diagnostics to emerging gene-based therapies, RNA molecules are now central to modern medicine. But as their use continues to grow, so does a fundamental challenge: producing RNA quickly, accurately and with the flexibility needed for next-generation biomedical applications.
Scientists at the University of California, Irvine have now taken a major step toward solving that problem.
In a study recently published in Nature Chemical Biology, a research team led by John Chaput, UC Irvine professor of pharmaceutical sciences, reports the creation of a powerful new enzyme that efficiently synthesizes RNA – something no natural DNA-copying enzyme can do. The engineered enzyme, known as C28, produces RNA at near-natural speeds while maintaining high accuracy and the ability to copy long sequences.
“DNA polymerases are naturally designed to reject RNA,” Chaput said. “What surprised us is that we were able to overcome this barrier not by redesigning the enzyme’s active site, but by letting evolution find unexpected structural solutions.”
Rather than manually engineering the enzyme, the researchers turned to directed evolution. Using a high-throughput, single-cell screening platform, the team recombined related polymerase genes and tested millions of enzyme variants in parallel. After only a few rounds of selection, they identified C28 – a polymerase carrying dozens of mutations spread throughout the protein that collectively enable efficient RNA synthesis.
The result is an enzyme with unusual versatility. In addition to synthesizing RNA, C28 can perform reverse transcription, copying RNA back into DNA, and can generate hybrid DNA-RNA molecules using standard polymerase chain reaction techniques. The enzyme also readily accepts several chemically modified RNA building blocks, including those used in mRNA vaccines and RNA-based therapeutics.
This combination of speed, accuracy and flexibility could make C28 a valuable new tool for researchers and biotechnology developers, particularly in applications that require customized or chemically modified RNA molecules.
Beyond its practical applications, the research underscores the power of directed evolution to create entirely new molecular functions – capabilities that do not exist in nature but can be unlocked through carefully designed selection strategies.
“This work shows that enzymes are far more adaptable than we once thought,” Chaput said. “By harnessing evolution, we can create new molecular tools that open the door to advances in RNA biology, synthetic biology and biomedical innovation.”
Other UC Irvine team members were Esau Medina, Victoria Maola Gross, Mohammad Hajjar, Ethan Ho, Alexandria Horton, Nicholas Chim and Grace Ko.
Note: Modified RNA (modRNA) is a specific type of messenger RNA (mRNA) that has been chemically altered to improve its function. While all modRNA is mRNA, not all mRNA is modified.
Funding: The National Science Foundation supported the research.
Published in journal: Nature Chemical Biology
Title: Rapid evolution of a highly efficient RNA polymerase by homologous recombination
Authors: Esau L. Medina, Victoria A. Maola, Mohammad Hajjar, Grace K. Ko, Ethan J. Ho, Alexandria R. Horton, Nicholas Chim, and John C. Chaput
Source/Credit: University of California, Irvine
Reference Number: bchm021026_01
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