Determination of 3’–5’ exonuclease and protein histidine kinase activity for selected NM23-H1 mutant variants. (a) Purified preparations of wild-type or mutant NM23-H1 proteins (500 ng) were incubated with 10 fmol of a single-stranded, 5’-radiolabeled DNA substrate for the indicated times at room temperature, as described in Materials and Methods. Extent of DNA cleavage was analyzed by electrophoresis through denaturing 10% polyacrylamide gels and visualization by phosphorimaging. P, radiolabeled probe. (b) 3’–5’ exonuclease assay of E5A mutant is shown. C, Quantitation of results in panels A and B are shown, expressed as total nucleotides (nt) cleaved. (c) Protein histidine kinase activity was analyzed for wild-type and mutant variants of NM23-H1. NM23-H1 proteins were auto-radiolabeled with [32P-γ] ATP, followed by incubation with unlabeled NM23-H2 substrate for the indicated times.