Clinical use of CRISPR-based gene editing has mostly involved modifying cells outside the body before reintroducing them. A key challenge has been that the most reliable gene-editing tools exceed the size limits of viral delivery systems needed to reach specific cells or tissues inside patients.
A University of Texas at Austin team, working with Metagenomi Therapeutics, created a smaller CRISPR nuclease called Al3Cas12f. This naturally occurring bacterial enzyme fits within adeno-associated virus vectors while maintaining strong gene-editing performance. The researchers published their findings in Nature Structural & Molecular Biology.
Metagenomi identified Al3Cas12f for its compact size and efficiency. UT Austin scientists then analyzed and refined it, producing an enhanced version that worked better in human cells. Study co-author David Taylor, said, "We uncovered mechanistic features that explain why some Cas12f enzymes are more efficient than others." He added that this knowledge allows for rational design of improved variants that retain a small size suitable for delivery. Taylor also noted, "Importantly, we also identified Al3Cas12f as a highly efficient nuclease across multiple genomic targets, making it a strong candidate for future therapeutic development."
AAV vectors can carry instructions for about 1,000 amino acids, but top nucleases are too large. This has restricted CRISPR applications to accessible tissues like blood and bone marrow. Smaller Cas12f nucleases (400-700 amino acids) had potential but underperformed in human cells until this study.
The team found Al3Cas12f outperformed two other Cas12f enzymes previously used in mice to edit muscular dystrophy genes. Using cryo-electron microscopy and machine learning, they modeled its structure and DNA interactions. Al3Cas12f showed a larger interface between components for greater stability. Taylor explained, "The expanded interface means the enzyme is much more stable. Compared to the others we looked at, Al3Cas12f basically comes preassembled and ready to go shortly after its pieces are produced."
The researchers engineered variants, with Al3Cas12f RKK achieving over 80% efficiency in human leukemia cells. They targeted genes linked to cancer, atherosclerosis and ALS.
Next steps include testing RKK in AAV vectors. The work was supported by NIH's National Institute of General Medical Sciences. Erica Brown, Acting NIGMS Director, said, "Smart delivery of gene-editing systems is a powerful notion with broad clinical implications, and this basic science finding takes us a significant step toward that future."
Publication details: Guan, K., Ocampo, R.F., Matheus Carnevali, P.B. et al. Comparative characterization of Cas12f orthologs reveals mechanistic features underlying enhanced genome editing efficiency. Nat Struct Mol Biol (2026). https://doi.org/10.1038/s41594-026-01788-6
A University of Texas at Austin team, working with Metagenomi Therapeutics, created a smaller CRISPR nuclease called Al3Cas12f. This naturally occurring bacterial enzyme fits within adeno-associated virus vectors while maintaining strong gene-editing performance. The researchers published their findings in Nature Structural & Molecular Biology.
Metagenomi identified Al3Cas12f for its compact size and efficiency. UT Austin scientists then analyzed and refined it, producing an enhanced version that worked better in human cells. Study co-author David Taylor, said, "We uncovered mechanistic features that explain why some Cas12f enzymes are more efficient than others." He added that this knowledge allows for rational design of improved variants that retain a small size suitable for delivery. Taylor also noted, "Importantly, we also identified Al3Cas12f as a highly efficient nuclease across multiple genomic targets, making it a strong candidate for future therapeutic development."
AAV vectors can carry instructions for about 1,000 amino acids, but top nucleases are too large. This has restricted CRISPR applications to accessible tissues like blood and bone marrow. Smaller Cas12f nucleases (400-700 amino acids) had potential but underperformed in human cells until this study.
The team found Al3Cas12f outperformed two other Cas12f enzymes previously used in mice to edit muscular dystrophy genes. Using cryo-electron microscopy and machine learning, they modeled its structure and DNA interactions. Al3Cas12f showed a larger interface between components for greater stability. Taylor explained, "The expanded interface means the enzyme is much more stable. Compared to the others we looked at, Al3Cas12f basically comes preassembled and ready to go shortly after its pieces are produced."
The researchers engineered variants, with Al3Cas12f RKK achieving over 80% efficiency in human leukemia cells. They targeted genes linked to cancer, atherosclerosis and ALS.
Next steps include testing RKK in AAV vectors. The work was supported by NIH's National Institute of General Medical Sciences. Erica Brown, Acting NIGMS Director, said, "Smart delivery of gene-editing systems is a powerful notion with broad clinical implications, and this basic science finding takes us a significant step toward that future."
Publication details: Guan, K., Ocampo, R.F., Matheus Carnevali, P.B. et al. Comparative characterization of Cas12f orthologs reveals mechanistic features underlying enhanced genome editing efficiency. Nat Struct Mol Biol (2026). https://doi.org/10.1038/s41594-026-01788-6