Kinetic mechanism for single-stranded DNA binding and translocation by Saccharomyces cerevisiae Isw2.
The chromatin remodeling complex Isw2 from Saccharomyces cerevisiae (yIsw2) mobilizes nucleosomes through an ATP-dependent reaction that is coupled to the translocation of the helicase domain of the enzyme along intranucleosomal DNA. In this study, we demonstrate that yIsw2 is capable of translocating along single-stranded DNA in a reaction that is coupled to ATP hydrolysis. We propose that single-stranded DNA translocation by yIsw2 occurs through a series of repeating uniform steps with an overall macroscopic processivity (P) of 0.90 +/- 0.02, corresponding to an average translocation distance of 20 +/- 2 nucleotides before dissociation. This processivity corresponds well to the processivity of nucleosome sliding by yIsw2, thus arguing that single-stranded DNA translocation or tracking may be fundamental to the double-stranded DNA translocation required for effective nucleosome mobilization. Furthermore, we find evidence that a slow initiation process, following DNA binding, may be required to make yIsw2 competent for DNA translocation. We also provide evidence that this slow initiation process may correspond to the second step of a two-step DNA binding mechanism by yIsw2 and a quantitative description of the kinetics of this DNA binding mechanism.
Fischer
Christopher J
CJ
Department of Physics and Astronomy, University of Kansas, 1251 Wescoe Hall Drive, 1082 Malott Hall, Lawrence, Kansas 66045, USA. shark@ku.edu
Yamada
Kazuhiro
K
Fitzgerald
Daniel J
DJ
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000494:pharmacology; D004277:DNA, Single-Stranded / Q000378:metabolism*; D006868:Hydrolysis / Q000187:drug effects; D007700:Kinetics; D000226:Mitochondrial ADP, ATP Translocases / Q000378:metabolism; D008958:Models, Molecular; D011485:Protein Binding / Q000187:drug effects; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000201:enzymology*; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism*
D004277:DNA, Single-Stranded; C086468:ISWI protein; D014157:Transcription Factors; D000255:Adenosine Triphosphate; D000226:Mitochondrial ADP, ATP Translocases; D000251:Adenosine Triphosphatases