Requirements for ATP binding and hydrolysis in RecA function in Escherichia coli.
RecA is essential for recombination, DNA repair and SOS induction in Escherichia coli. ATP hydrolysis is known to be important for RecA's roles in recombination and DNA repair. In vitro reactions modelling SOS induction minimally require ssDNA and non-hydrolyzable ATP analogues. This predicts that ATP hydrolysis will not be required for SOS induction in vivo. The requirement of ATP binding and hydrolysis for SOS induction in vivo is tested here through the study of recA4159 (K72A) and recA2201 (K72R). RecA4159 is thought to have reduced affinity for ATP. RecA2201 binds, but does not hydrolyse ATP. Neither mutant was able to induce SOS expression after UV irradiation. RecA2201, unlike RecA4159, could form filaments on DNA and storage structures as measured with RecA-GFP. RecA2201 was able to form hybrid filaments and storage structures and was either recessive or dominant to RecA(+), depending on the ratio of the two proteins. RecA4159 was unable to enter RecA(+) filaments on DNA or storage structures and was recessive to RecA(+). It is concluded that ATP hydrolysis is essential for SOS induction. It is proposed that ATP binding is essential for storage structure formation and ability to interact with other RecA proteins in a filament.
Renzette
Nicholas
N
Molecular and Cellular Biology Graduate Program, Morrill Science Center, University of Massachusetts at Amherst, Amherst, MA 01003, USA.
Sandler
Steven J
SJ
D000255:Adenosine Triphosphate / Q000378:metabolism*; D002876:Chromosomes, Bacterial; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism* / Q000528:radiation effects; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D006868:Hydrolysis; D009154:Mutation; D011485:Protein Binding; D011938:Rec A Recombinases / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D014466:Ultraviolet Rays
D029968:Escherichia coli Proteins; D000255:Adenosine Triphosphate; D011938:Rec A Recombinases