APOBEC1 Mutations Reduce Off-Target SNVs in Base Editing

The patent applies parameter changes by modifying the amino acid sequence of the APOBEC1 enzyme (changing specific residues such as R126E, W90Y, R90E mutations). These parameter changes in the enzyme's primary structure alter its DNA binding affinity and catalytic activity, enabling it to maintain high editing efficiency at the target site while reducing off-target single-nucleotide mutations. This directly resolves the technical contradiction by tuning the enzyme's parameters to achieve selective activity.

The patent applies local quality by creating spatially differentiated effects of the modified APOBEC1 enzyme. The enzyme exhibits high catalytic activity at the intended target site (local high efficiency) while showing reduced activity at off-target sites (local low mutation rate). This is achieved through mutations that fine-tune the enzyme's DNA recognition and binding properties, allowing it to discriminate between target and non-target sequences, thus resolving the contradiction between efficiency and off-target effects.

The patent modifies parameters of the cytosine deaminase (APOBEC1) by introducing specific amino acid mutations (R126E, W90Y, R90E) that alter its DNA binding characteristics. These parameter changes reduce the enzyme's non-specific DNA binding affinity while preserving its catalytic function at the target site, thereby maintaining reliable editing activity while reducing off-target effects through precise parameter optimization.

The patent applies inversion by reversing the conventional approach: instead of strengthening DNA binding to improve editing function, the patent weakens the overall DNA binding affinity through mutations and compensates by enhancing specificity at the target site. This inverted strategy reduces off-target binding while maintaining on-target efficiency, resolving the contradiction between function and off-target effects.