FIG. 2.

Specificity of processing by integrases with serine-124 substitutions. (A) Autoradiogram of processing assay. Double-stranded 18-mers derived from the U3 end of RSV DNA were 5′ labeled on the strand that contains the conserved CA and incubated with protein buffer (lanes 1 and 8), wild-type RSV integrase (lanes 2 and 9), or RSV integrases with the indicated amino acid substitutions. Reactions 1 to 7 used 10 mM Mn²⁺, and reactions 8 to 14 used 5 mM Mg²⁺. Reactions were conducted and analyzed as in Materials and Methods. Nucleotide sizes, the 18-mer substrate, and the 16-mer and 15-mer products are indicated (reflecting nicks at the −2 and −3 sites, respectively). (B) Quantitation of replicate reactions. The ratios of 16-mer to 15-mer products were calculated for triplicate experiments as described in Materials and Methods. The bars are aligned with the corresponding lane from the autoradiograms in panel A. Note that the right graph has a larger scale than the left graph.