ABSTRACT
BACKGROUND: Tangle analysis has been applied successfully to study proteins which bind two segments of DNA and can knot and link circular DNA. We show how tangle analysis can be extended to model any stable protein-DNA complex. RESULTS: We discuss a computational method for finding the topological conformation of DNA bound within a protein complex. We use an elementary invariant from knot theory called colorability to encode and search for possible DNA conformations. We apply this method to analyze the experimental results of Pathania, Jayaram, and Harshey (Cell 2002). We show that the only topological DNA conformation bound by Mu transposase which is biologically likely is the five crossing solution found by Pathania et al (although other possibilities are discussed). CONCLUSION: Our algorithm can be used to analyze the results of the experimental technique described in Pathania et al in order to determine the topological conformation of DNA bound within a stable protein-DNA complex.
Subject(s)
Bacteriophage mu/genetics , DNA Transposable Elements/genetics , DNA, Superhelical/chemistry , DNA, Viral/chemistry , DNA-Binding Proteins/chemistry , Software , Transposases/chemistry , Algorithms , Bacteriophage mu/metabolism , Binding Sites , DNA, Superhelical/genetics , DNA, Viral/genetics , DNA-Binding Proteins/metabolism , Integrases/chemistry , Integrases/metabolism , Models, Chemical , Models, Genetic , Nucleic Acid Conformation , Protein Binding , Protein Conformation , Transposases/metabolismABSTRACT
The Flp recombinase of yeast and the Cre recombinase of bacteriophage P1 both belong to the lambda-integrase (Int) family of site-specific recombinases. These recombination systems recognize recombination-target sequences that consist of two 13bp inverted repeats flanking a 6 or 8bp spacer sequence. Recombination reactions involve particular geometric and topological relationships between DNA target sites at synapsis, which we investigate using nicked-circular DNA molecules. Examination of the tertiary structure of synaptic complexes formed on nicked plasmid DNAs by atomic-force microscopy, in conjunction with detailed topological analysis using the mathematics of tangles, shows that only a limited number of recombination-site topologies are consistent with the global structures of plasmids bearing directly and inversely repeated sites. The tangle solutions imply that there is significant distortion of the Holliday-junction intermediate relative to the planar structure of the four-way DNA junction present in the Flp and Cre co-crystal structures. Based on simulations of nucleoprotein structures that connect the two-dimensional tangle solutions with three-dimensional models of the complexes, we propose a recombination mechanism in which the synaptic intermediate is characterized by a non-planar, possibly near-tetrahedral, Holliday-junction intermediate. Only modest conformational changes within this structure are needed to form the symmetric, planar DNA junction, which may be characteristic of shorter-lived intermediates along the recombination pathway.
