Allosteric interactions of DNA and nucleotides with S. cerevisiae RSC.

Shuja Shafi Malik, Evan Rich, Ramya Viswanathan, Bradley R Cairns, Christopher J Fischer

Department of Physics and Astronomy, University of Kansas, 1251 Wescoe Hall Dr., 1082 Malott Hall, Lawrence, Kansas 66045, USA.

Biochemistry 2011 Sep 20

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RSC (remodel the structure of chromatin) is an essential chromatin remodeler of Saccharomyces cerevisiae that has been shown to have DNA translocase properties. We studied the DNA binding properties of a "trimeric minimal RSC" (RSCt) of the RSC chromatin remodeling complex and the effect of nucleotides on this interaction using fluorescence anisotropy. RSCt binds to 20 bp fluorescein-labeled double-stranded DNA with a K(d) of ∼100 nM. The affinity of RSCt for DNA is reduced in the presence of AMP-PNP and ADP in a concentration-dependent manner with the addition of AMP-PNP having more pronounced effect. These differences in the magnitude at which the binding of ADP and AMP-PNP affects the affinity of DNA binding by RSCt suggest that the physical movement of the enzyme along DNA begins between the binding of ATP and its subsequent hydrolysis. Furthermore, the fact that the highest affinity for DNA binding by RSCt occurs in the absence of bound nucleotide offers a mechanistic explanation for the apparent low processivity of DNA translocation by the enzyme. © 2011 American Chemical Society

Citation

Shuja Shafi Malik, Evan Rich, Ramya Viswanathan, Bradley R Cairns, Christopher J Fischer. Allosteric interactions of DNA and nucleotides with S. cerevisiae RSC. Biochemistry. 2011 Sep 20;50(37):7881-90