FIG. 4.

Effects of temperature (A), pH (B), divalent cations Mg²⁺ and Mn²⁺ (C), and salt concentration (D) on purified AcMNPV alkaline exonuclease activity. For these experiments, single-stranded DNA (2 μg of labeled DNA plus 4 μg of salmon sperm DNA) was mixed with 10 ng of purified AN. Samples were prepared for each condition tested and then incubated for 20 min. Standard conditions were 37°C, 50 mM Tris (pH 9.0), 5 mM MgCl, and 0.1 mg of BSA per ml in 200 μl unless otherwise indicated. The digested DNA was mixed with 0.25 mg of BSA, then precipitated with 5% trichloroacetic acid, and centrifuged for 5 min in a microcentrifuge; the supernatants were mixed with 3.5 ml of scintillation cocktail (formula A-989; Packard) and counted. All samples were done in triplicate, and each point represents an average of the values. Very little deviation from the average was observed.