Prime editing is a powerful genome engineering technique that allows for precise single-base substitutions and small insertions and deletions without requiring homologous recombination or double-strand DNA breaks in eukaryotic cells. However, its application in bacteria has been limited due to the presence of factors that impede efficient prime editing. This study identifies *sbcB*, a 3′-5′ DNA exonuclease, as a key genetic determinant that hinders prime editing in *Escherichia coli*. Combinatorial deletions of *sbcB* with two additional 3′-5′ DNA exonucleases, *xseA* and *exoX*, significantly enhance prime editing efficiency by up to 100-fold. The authors propose a 3′-directed hydrolysis model where these exonucleases degrade prime editing intermediates, reducing editing efficiency. They also demonstrate that inhibiting these exonucleases via CRISPRi can achieve efficient prime editing in wild-type *E. coli*. This work paves the way for versatile applications of prime editing in bacterial genome engineering.Prime editing is a powerful genome engineering technique that allows for precise single-base substitutions and small insertions and deletions without requiring homologous recombination or double-strand DNA breaks in eukaryotic cells. However, its application in bacteria has been limited due to the presence of factors that impede efficient prime editing. This study identifies *sbcB*, a 3′-5′ DNA exonuclease, as a key genetic determinant that hinders prime editing in *Escherichia coli*. Combinatorial deletions of *sbcB* with two additional 3′-5′ DNA exonucleases, *xseA* and *exoX*, significantly enhance prime editing efficiency by up to 100-fold. The authors propose a 3′-directed hydrolysis model where these exonucleases degrade prime editing intermediates, reducing editing efficiency. They also demonstrate that inhibiting these exonucleases via CRISPRi can achieve efficient prime editing in wild-type *E. coli*. This work paves the way for versatile applications of prime editing in bacterial genome engineering.