Inhibition of recA protein promoted ATP hydrolysis. 2. Longitudinal assembly and disassembly of recA protein filaments mediated by ATP and ADP.
There are at least two major conformations of recA nucleoprotein filaments formed on poly-(deoxythymidylic acid) [poly(dT)], one stabilized by ATP [or adenosine 5'-O-(3-thiotriphosphate) (ATP gamma S)] and one stabilized by ADP. Assembly of filaments in the ATP conformation is much faster than assembly in the ADP conformation. A third conformation may be present in the absence of nucleotides. The ATP and ADP conformations are mutually exclusive. When a mixture of ATP and ADP is present, recA protein binding is a function of the ADP/ATP ratio. Complete dissociation is observed when the ratio becomes 1.0-1.5. When a mixture of ATP and ADP is present at the beginning of a reaction, a transient phase lasting several minutes is observed in which the system approaches the state characteristic of the new ADP/ATP ratio. This phase is manifested by a lag in ATP hydrolysis when ATP is added to preformed ADP filaments, and by a burst in ATP hydrolysis in all other cases. More than 15 ATPs are hydrolyzed per bound recA monomer during the burst phase. The transient phase reflects an end-dependent disassembly process propagated longitudinally through the filament, rather than a slow conformation change in individual recA monomers or a slow exchange of one nucleotide for the other. The hysteresis exhibited by the system provides a number of insights relevant to the mechanism of recA-mediated DNA strand exchange.
Lee
J W
JW
Department of Biochemistry, College of Agricultural and Life Sciences, University of Wisconsin-Madison 53706.
Cox
M M
MM
D000244:Adenosine Diphosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism*; D006868:Hydrolysis; D007700:Kinetics; D011938:Rec A Recombinases / Q000378:metabolism*
C022571:adenosine 5'-O-(3-thiotriphosphate); D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D011938:Rec A Recombinases