Stimulation of the Mu DNA strand cleavage and intramolecular strand transfer reactions by the Mu B protein is independent of stable binding of the Mu B protein to DNA.
Interactions between the Mu A and Mu B proteins are important in the early steps of the in vitro transposition of a mini-Mu plasmid. We have examined these interactions by assaying Mu B stimulation of Mu A-mediated strand cleavage and strand transfer reactions. We have previously shown that in the presence of ATP the Mu B protein can stimulate the Mu A-directed cleavage reaction of mini-Mu plasmids carrying a terminal base pair mutation (Surette, M.G., Harkness, T., and Chaconas, G. (1991) J. Biol. Chem. 266, 3118-3124). Here we demonstrate that in the absence of a non-Mu DNA target molecule the Mu B protein stimulates intramolecular integration of a mini-Mu in an ATP-dependent fashion. Furthermore, modification of the Mu B protein with N-ethylmaleimide severely compromises the ability of B to form a stable complex with DNA; however, the modified protein stimulates the strand cleavage and intramolecular strand transfer reactions as efficiently as the untreated protein. These results indicate that the Mu B protein is capable of stimulating the Mu A protein through direct interaction in the absence of stable Mu B-DNA complex formation. Our results increase the spectrum of Mu B protein activities and uncouple the stimulatory properties of the Mu B protein from stable DNA binding but not the ATP cofactor requirement.
Surette
M G
MG
Department of Biochemistry, University of Western Ontario, London, Canada.
Chaconas
G
G
D000251:Adenosine Triphosphatases / Q000378:metabolism; D010583:Bacteriophage mu / Q000235:genetics / Q000378:metabolism*; D004279:DNA, Viral / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004587:Electrophoresis, Agar Gel; D005033:Ethylmaleimide / Q000494:pharmacology; D007700:Kinetics; D009713:Nucleotidyltransferases / Q000378:metabolism; D019895:Transposases; D014764:Viral Proteins / Q000378:metabolism*
D004279:DNA, Viral; D004268:DNA-Binding Proteins; C050938:Mu B protein, bacteriophage; D014764:Viral Proteins; D009713:Nucleotidyltransferases; D019895:Transposases; D000251:Adenosine Triphosphatases; D005033:Ethylmaleimide