Figure 1

KRC fusion proteins and site-selection EMSA Figure 1A. KRC fusion proteins. (Top) The full-length KRC protein is described schematically. In the ZAS-N and ZAS-C DNA-binding domains the z inc-fingers, a cidic regions, and serine-threonine-rich regions highlighted. ZASN, ZAS-N domain; ZF3, zinc finger 3; NLS, nuclear localization signal; GTP, GTPase motif; ZASC, ZAS-C motif. (Bottom) KRC fusion proteins, KRC/ZAS-N and KRC/ZAS-C are described schematically. KRC/ZAS-N is a S-tag fusion protein containing the ZAS-N DNA-binding domain (nt 949–2167) KRC/ZAS-C is an Mbp fusion protein containing the ZAS-C DNA-binding domain (nt 5544–7015). These are the fusion proteins used in the site-selection assay described in this paper. Figure 1B and 1C. Electrophoretic mobility shift assays of the site selection procedures. (Bottom) A portion of the oligonucleotides (~0.2 ng and 5000 cpm) recovered from each round of site selection was 32P-labeled and incubated with KRC fusion proteins (~0.5 μg), (B) KRC/ZAS-N and (C) KRC/ZAS-C, in the presence of 10 μg poly(dI-dC). DNA-protein complexes and free probes were resolved in 6% polyacrylamide gels and visualized by exposing dried gels to X-ray films. The probes used in lanes 1 through 5 were derived from aliquots of DNA recovered from round one through five of site selection, respectively. C, DNA-protein complexes; and F, free probes.