Figure 4.

Acceptor nucleotide for VP4 methyltransferase activity. (A) Recombinant VP4 was incubated with cold GTP in the presence of 2 μM Ado[methyl-³H]Met (87 Ci/mmol), 50 mM Tris⋅HCl (pH 7.9), 9 mM MgCl₂, and 6 mM DTT. After incubation for 2 hr at 30°C and treatment with proteinase K, the reaction products were extracted with phenol/chloroform and precipitated with ethanol. Samples were analyzed by HPLC. The positions of unincorporated AdoMet and m⁷GpppG are indicated. (B) Purified CLPs encapsidating VP4 (30 μg) were incubated with 6 μM ([α-³²P]GTP (400 Ci/mmol) and 2 μM AdoMet in 50 mM Tris⋅HCl (pH 7.9), 9 mM MgCl₂, and 6 mM DTT. Samples were analyzed as described. Position of m⁷GpppGm is indicated.