TY - JOUR
T1 - PcrA helicase dismantles RecA filaments by reeling in DNA in uniform steps
AU - Park, Jeehae
AU - Myong, Sua
AU - Niedziela-Majka, Anita
AU - Lee, Kyung Suk
AU - Yu, Jin
AU - Lohman, Timothy M.
AU - Ha, Taekjip
N1 - Funding Information:
We thank C. Joo, R. Roy, M. Schlierf, S. Syed, M.C. McKinney, M.K. Nahas, B. Okumus, S.A. McKinney, E. Bozek, and J.D. Smith for experimental help; S. Arslan, J. Yodh, H. Balci, I. Cisse, B.C. Stevens, G. Lee, M. Brenner, E. Tomko, C. Fischer, and M. Spies for helpful discussion; I. Cann and his lab for providing Staphylococcus aureus PcrA for the initial studies; R. Pugh for help on ExoIII footprint assay; M. Schlierf, J. Yodh, H. Balci, and E. Tomko for careful reading of the manuscript. Funds were provided by the National Institutes of Health (GM-065367 to T.H. and GM-045948 to T.M.L.) and by the National Science Foundation (0822613 and 0646550 to T.H.). T.H. is an investigator with the Howard Hughes Medical Institute.
PY - 2010/8
Y1 - 2010/8
N2 - Translocation of helicase-like proteins on nucleic acids underlies key cellular functions. However, it is still unclear how translocation can drive removal of DNA-bound proteins, and basic properties like the elementary step size remain controversial. Using single-molecule fluorescence analysis on a prototypical superfamily 1 helicase, Bacillus stearothermophilus PcrA, we discovered that PcrA preferentially translocates on the DNA lagging strand instead of unwinding the template duplex. PcrA anchors itself to the template duplex using the 2B subdomain and reels in the lagging strand, extruding a single-stranded loop. Static disorder limited previous ensemble studies of a PcrA stepping mechanism. Here, highly repetitive looping revealed that PcrA translocates in uniform steps of 1 nt. This reeling-in activity requires the open conformation of PcrA and can rapidly dismantle a preformed RecA filament even at low PcrA concentrations, suggesting a mode of action for eliminating potentially deleterious recombination intermediates.
AB - Translocation of helicase-like proteins on nucleic acids underlies key cellular functions. However, it is still unclear how translocation can drive removal of DNA-bound proteins, and basic properties like the elementary step size remain controversial. Using single-molecule fluorescence analysis on a prototypical superfamily 1 helicase, Bacillus stearothermophilus PcrA, we discovered that PcrA preferentially translocates on the DNA lagging strand instead of unwinding the template duplex. PcrA anchors itself to the template duplex using the 2B subdomain and reels in the lagging strand, extruding a single-stranded loop. Static disorder limited previous ensemble studies of a PcrA stepping mechanism. Here, highly repetitive looping revealed that PcrA translocates in uniform steps of 1 nt. This reeling-in activity requires the open conformation of PcrA and can rapidly dismantle a preformed RecA filament even at low PcrA concentrations, suggesting a mode of action for eliminating potentially deleterious recombination intermediates.
KW - DNA
UR - http://www.scopus.com/inward/record.url?scp=77955605175&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=77955605175&partnerID=8YFLogxK
U2 - 10.1016/j.cell.2010.07.016
DO - 10.1016/j.cell.2010.07.016
M3 - Article
C2 - 20723756
AN - SCOPUS:77955605175
SN - 0092-8674
VL - 142
SP - 544
EP - 555
JO - Cell
JF - Cell
IS - 4
ER -