Abstract
Serine integrases catalyse site-specific recombination to integrate and excise bacteriophage genomes into and out of their host's genome. These enzymes exhibit remarkable directionality; in the presence of the integrase alone, recombination between attP and attB DNA sites is efficient and irreversible, giving attL and attR products which do not recombine further. However, in the presence of the bacteriophage-encoded recombination directionality factor (RDF), integrase efficiently promotes recombination between attL and attR to re-form attP and attB. The DNA substrates and products of both reactions are approximately isoenergetic, and no cofactors (such as adenosine triphosphate) are required for recombination. The thermodynamic driving force for directionality of these reactions is thus enigmatic. Here, we present a minimal mathematical model which can explain the directionality and regulation of both ‘forward’ and ‘reverse’ reactions. In this model, the substrates of the ‘forbidden’ reactions (between attL and attR in the absence of RDF, attP and attB in the presence of RDF) are trapped as inactive protein–DNA complexes, ensuring that these ‘forbidden’ reactions are extremely slow. The model is in good agreement with the observed in vitro kinetics of recombination by ϕC31 integrase, and defines core features of the system necessary and sufficient for directionality.
Highlights
Serine integrases catalyse integration of a circular bacteriophage genomic DNA molecule into the bacterial host chromosomal DNA, by recombination between an attP site in the phage DNA and an attB site in the host DNA
recombination directionality factor (RDF) interacts with integrase and alters its properties so that it recombines the attL and attR sites to release the circular phage genomic DNA with an attP site, and leave an attB site in the host genome. (Hereafter, we refer to recombination between attP and attB as P Â B recombination, and recombination between attL and attR as L Â R recombination.)
The simulations with Model M quantitatively describe the key features of the data, such as the observed sharp stimulation of the L Â R(þR) reaction and inhibition of the P Â B(2R) reaction when the concentration of RDF reaches that of integrase
Summary
Serine integrases catalyse integration of a circular bacteriophage genomic DNA molecule into the bacterial host chromosomal DNA, by recombination between an attP site in the phage DNA and an attB site in the host DNA. We recently presented a detailed mathematical model of recombination by fC31 integrase (the first serine integrase to be identified, and the best-characterized to date) [5,7,8], which aimed to account as far as possible for the available biochemical, molecular and structural data [1] (here called ‘Model A’). We were motivated to create a highly simplified datadriven mathematical model of serine integrase-mediated recombination Such a minimal model with a simple structure and minimal number of parameters should be useful in analysis of the key principles of unidirectional reversible genetic transformations, which might be applicable to other biological systems. Owing to its simplicity, this model is more generally applicable and is adaptable to other integrase-mediated recombination systems
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