title: The same two monomers within a MuA tetramer provide the DDE domains for the strand cleavage and strand transfer steps of transposition.
abstract: The chemistry of Mu transposition is executed within a tetrameric form of the Mu transposase (MuA protein). A triad of DDE (Asp, Asp35Glu motif) residues in the central domain of MuA (DDE domain) is essential for both the strand cleavage and strand transfer steps of transposition. Previous studies had suggested that complete Mu transposition requires all four subunits in the MuA tetramer to carry an active DDE domain. Using a mixture of MuA proteins with either wild-type or altered att-DNA binding specificities, we have now designed specific arrangements of MuA subunits carrying the DDE domain. From analysis of the abilities of oriented tetramers to carry out DNA cleavage and strand transfer from supercoiled DNA, a new picture of the disposition of DNA and protein partners during transposition has emerged. For DNA cleavage, two subunits of MuA located at attL1 and attR1 (sites that undergo cleavage) provide DDE residues in trans. The same two subunits contribute DDE residues for strand transfer, also in trans. Thus, only two active DDE+ monomers within the tetramer carry out complete Mu transposition. We also show that when the attR1-R2 arrangement used on supercoiled substrates is tested for cleavage on linear substrates, alternative chemically competent DNA-protein associations are produced, wherein the functional DDE subunits are positioned at R2 rather than at R1.
authors.affiliation: Department of Microbiology, University of Texas, Austin, TX 78712, USA.
authors.forename: S Y
authors.initials: SY
authors.lastname: Namgoong
authors.forename: R M
authors.initials: RM
authors.lastname: Harshey
mesh_terms: D010583:Bacteriophage mu / Q000235:genetics*; D001665:Binding Sites; D004278:DNA, Superhelical / Q000378:metabolism; D004279:DNA, Viral; D011995:Recombination, Genetic; D013379:Substrate Specificity; D019895:Transposases
chemical_list: D004278:DNA, Superhelical; D004279:DNA, Viral; D019895:Transposases