A second high affinity HU binding site in the phage Mu transpososome.
The bacteriophage Mu in vitro transposition reaction proceeds through several higher order nucleoprotein intermediates (transpososomes). One of the requirements for complex assembly is the Escherichia coli sequence-independent DNA-binding protein, HU. This protein has an affinity for Mu transpososomes, which is at least 100 times greater than for supercoiled DNA (Lavoie and Chaconas, 1990). We have recently identified one such high affinity binding site at the Mu left end by converting HU into a chemical nuclease (Lavoie and Chaconas, 1993). Using immunoelectron microscopy, we now report high affinity HU binding to a region(s) distinct from the previously characterized left end site. HU can be removed from this region by a 0.5 M NaCl wash and subsequently reassembled into the complex with high efficiency. Furthermore, chemical modification of the Mu A protein in the Type 1 complex does not block HU reassembly into the transpososome; the high affinity HU binding observed is therefore unlikely to result from A-HU interactions. These findings are corroborated by the ability of eukaryotic HMG-1 to functionally replace HU in transpososome formation and to efficiently assemble into HU-depleted complexes. We propose that HU recognition of an altered DNA structure, rather than protein-protein interactions, mediates high affinity HU binding to Mu transpososomes.
Lavoie
B D
BD
Department of Biochemistry, University of Western Ontario, London, Canada.
Chaconas
G
G
D001426:Bacterial Proteins / Q000378:metabolism* / Q000648:ultrastructure; D010583:Bacteriophage mu / Q000378:metabolism*; D001665:Binding Sites; D004279:DNA, Viral / Q000378:metabolism* / Q000648:ultrastructure; D004268:DNA-Binding Proteins / Q000378:metabolism* / Q000648:ultrastructure; D004926:Escherichia coli / Q000378:metabolism*; D007700:Kinetics; D016253:Microscopy, Immunoelectron; D009690:Nucleic Acid Conformation; D009698:Nucleoproteins / Q000378:metabolism*; D017384:Sequence Deletion
D001426:Bacterial Proteins; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D009698:Nucleoproteins; C040330:histone-like protein HU, bacteria