Characterisation and reprogramming of bacteriophage mv4 integrase recombination specificity

Bacteriophage mv4 is a temperate bacterial virus able to integrate its genome at the 3-prime end of the tRNASER of Lactobacillus delbrueckii subsp. bulgaricus chromosome through site-specific recombination. Previous investigations revealed that the mv4Int/ attP / attB recombination module was atypical compared to conventional heterobivalent tyrosine recombinases, such as the paradigmatic Lambdavirus lambda integrase, suggesting alternative recombination mechanism. In vitro recombination assays with random DNA libraries were used to comprehensively delineate the mv4 recombination system. We showed that mv4Int is a 369-aa protein that exhibits all structural hallmarks of integrases from the Tn 916 family and interacts cooperatively with its recombination sites. We established that mv4Int distinguishes itself from classical heterobivalent integrases by a greater tolerance to nucleotide variations in attB and core- attP sites. We demonstrated that, upon considering nucleotide degeneracy, the 21-bp core- attP and attB recombination sites share structural similarities with classical heterobivalent integrase systems, with two 7-bp inverted-repeat regions corresponding to mv4Int core-binding sites surrounding a 7-bp strand-exchange region. Furthermore, our study highlighted compositional biases and nucleotide interdependencies within the core-binding regions that exerted a significant influence on the outcomes of recombination events. ### Competing Interest Statement The authors have declared no competing interest.