Site-specific recombinases as tools for heterologous gene integration

Abstract

Site-specific recombinases are the enzymes that catalyze site-specific recombination between two specific DNA sequences to mediate DNA integration, excision, resolution, or inversion and that play a pivotal role in the life cycles of many microorganisms including bacteria and bacteriophages. These enzymes are classified as tyrosine-type or serine-type recombinases based on whether a tyrosine or serine residue mediates catalysis. All known tyrosine-type recombinases catalyze the formation of a Holliday junction intermediate, whereas the catalytic mechanism of all known serine-type recombinases includes the 180° rotation and rejoining of cleaved substrate DNAs. Both recombinase families are further subdivided into two families; the tyrosine-type recombinases are subdivided by the recombination directionality, and the serine-type recombinases are subdivided by the protein size. Over more than two decades, many different site-specific recombinases have been applied to in vivo genome engineering, and some of them have been used successfully to mediate integration, deletion, or inversion in a wide variety of heterologous genomes, including those from bacteria to higher eukaryotes. Here, we review the recombination mechanisms of the best characterized recombinases in each site-specific recombinase family and recent advances in the application of these recombinases to genomic manipulation, especially manipulations involving site-specific gene integration into heterologous genomes.

Integrative and Conjugative Elements (ICEs): What They Do and How They Work

“…Frequently, in addition to the integrase, an ICE-encoded recombination directionality factor (RDF), sometimes called Xis, is required for excision. This factor influences the direction of recombination mediated by the recombinase (integrase) to favor excision (reviewed in 70). …”

Listeriaphages

“…40 Depending on the specific catalytic amino acid residue present, phage integrases are classified as serine or tyrosine recombinases. 41 Phage A118 features a serine integrase, which recognizes a sequence with only three basepairs homology between phage and bacterial attachment site. 13 In contrast, the PSA integrase is of the tyrosine type, and recognizes a 15 bp site located at the 3′ end of a tRNA Arg gene.…”

3.4 Rational Approaches for Transgene Expression: Targeted Integration and Episomal Maintenance

“…These recombinases play a pivotal role in the life cycles of many microorganisms including yeast, bacteria, and bacteriophages [67]. The recombinases can catalyze intra-and intermolecular reactions, namely DNA integration, excision, or inversion (see below).…”