The λ Integrase Site-specific Recombination Pathway

Abstract

The site-specific recombinase encoded by bacteriophage λ (Int) is responsible for integrating and excising the viral chromosome into and out of the chromosome of its Escherichia coli host. Int carries out a reaction that is highly directional, tightly regulated, and depends upon an ensemble of accessory DNA bending proteins acting on 240 bp of DNA encoding 16 protein binding sites. This additional complexity enables two pathways, integrative and excisive recombination, whose opposite, and effectively irreversible, directions are dictated by different physiological and environmental signals. Int recombinase is a heterobivalent DNA binding protein and each of the four Int protomers, within a multiprotein 400 kDa recombinogenic complex, is thought to bind and, with the aid of DNA bending proteins, bridge one arm- and one core-type DNA site. In the 12 years since the publication of the last review focused solely on the λ site-specific recombination pathway in Mobile DNA II, there has been a great deal of progress in elucidating the molecular details of this pathway. The most dramatic advances in our understanding of the reaction have been in the area of X-ray crystallography where protein-DNA structures have now been determined for of all of the DNA-protein interfaces driving the Int pathway. Building on this foundation of structures, it has been possible to derive models for the assembly of components that determine the regulatory apparatus in the P-arm, and for the overall architectures that define excisive and integrative recombinogenic complexes. The most fundamental additional mechanistic insights derive from the application of hexapeptide inhibitors and single molecule kinetics.

A bacteriophage mimic of the bacterial nucleoid-associated protein Fis

“…The remarkable similarities in NinH and Fis DNA binding and bending, mean that they have the potential to be functionally equivalent factors affecting several aspects of λ and E. coli biology. Fis fulfills an accessory role in excision of the λ prophage by manipulating attL and attR into an appropriate position for cleavage by integrase (Int) in complex with Xis bound to attR [6,7,11,56]. Fis binding at this site also contributes to phage integration [9][10][11].…”

“…Fis was originally identified due to its involvement in the topology manipulations necessary to drive site-specific inversion reactions involving the Hin and Gin recombinases [5]. Similarly, Fis works with Xis to direct excision of the λ prophage from the host chromosome [6][7][8] and can also stimulate phage integration reactions [9][10][11]. Fis condenses DNA by binding 21 bp segments via its non-specific DNA binding and bending activities [12].…”

Mechanistic modelling of a recombinase‐based two‐input temporal logic gate

“…The former have been shown to provide effective genetic switch mechanisms [11][12][13], but their functionality is often dependent on cell-specific cofactors, as in the case of l integrase [14]. This poses a similar problem to that of transcriptional systems with respect to deploying modules across multiple organisms.…”

Mechanistic Modeling of a Rewritable Recombinase Addressable Data Module

“…SSRs are classified as belonging to two groups, the tyrosine recombinases and the serine recombinases. The former have been shown to provide effective genetic switch mechanisms however, their functionality is often dependent upon cell-specific cofactors as in the case of λ integrase [8]. This is problematic in a similar vein to that of the transcriptional systems when looking to deploy modules across multiple organisms.…”