# Triple base editor catalyzes saturation mutation of adenine, cytidine, and guanine

## TL;DR

Scientists created a new base editor that can efficiently mutate three DNA bases, enabling broader genetic screening and therapeutic applications.

## Contribution

The development of a triple base editor (ACG-BEs) that enables saturation mutagenesis across adenine, cytosine, and guanine.

## Key findings

- ACG-BEs achieved up to 80.5% conversion efficiency for A-to-G/C/T in HEK293T cells.
- The editor enabled identification of novel mutations in the HBG1/2 promoter region for γ-globin activation.
- ACG-BEs offer enhanced DNA sequence diversity for genetic screening and therapeutic strategies.

## Abstract

Current base editors act on a maximum of two base substrates and generate limited base conversions or transversions, hindering their applicability for inducing DNA sequence diversity. Here, we engineered a triple base editor (named ACG-BEs) using a fusion of adenine base editor with high A/C catalytic activity and evolved N-methylpurine DNA glycosylase. ACG-BEs enables efficient, multiplexed saturation mutagenesis across adenine (A), cytosine (C), and guanine (G), achieving conversion efficiencies of up to 80.5% for A-to-G/C/T, 75.8% for C-to-T/G/A, and 63.4% for G-to-C/T/A in HEK293T cells. Leveraging ACG-BEs, we identify novel mutations in the HBG1/2 promoter region that confer efficient activation of γ-globin expression in HUDEP-2 cells—a promising advancement for therapeutic strategies targeting hemoglobinopathies. These findings highlight ACG-BEs as a cutting-edge platform for multiplexed saturation mutagenesis, offering broad applications in genetic screening and therapeutic base mutation introduction through enhanced DNA sequence diversity.

Graphical Abstract